The essential oil secretory structures of Prostanthera ovalifolia (Lamiaceae)

The structure of the essential oil secretory tissues of Prostanthera ovalifolia R.Br was investigated using bright- and dark-field optical microscopy, and scanning and transmission electron microscopy. The leaves of P. ovalifolia have glandular trichomes of the peltate type common to many Lamiaceae species. The trichomes consist of a basal cell embedded in the epidermis, a stalk cell with heavily cutinized walls and a 16-celled secretory head, but they differ from those of many previously reported Lamiaceae species in their morphological form defined by the elevated cuticle. The sub-cuticular space contains a mixture of lipid and aqueous phases. Secretory cells have dense cytoplasm with many leucoplasts present. Volatile terpenoids are eliminated from the cytoplasm into the sub-cuticular space, the site of essential oil accumulation, via granulocrine secretion.     

Organization of monoterpene biosynthesis in Mentha. Immunocytochemical localizations of geranyl diphosphate synthase, limonene-6-hydroxylase, isopiperitenol dehydrogenase, and pulegone reductase

We present immunocytochemical localizations of four enzymes involved in p-menthane monoterpene biosynthesis in mint: the large and small subunits of peppermint (Mentha x piperita) geranyl diphosphate synthase, spearmint (Mentha spicata) (-)-(4S)-limonene-6-hydroxylase, peppermint (-)-trans-isopiperitenol dehydrogenase, and peppermint (+)-pulegone reductase. All were localized to the secretory cells of peltate glandular trichomes with abundant labeling corresponding to the secretory phase of gland development. Immunogold labeling of geranyl diphosphate synthase occurred within secretory cell leucoplasts, (-)-4S-limonene-6-hydroxylase labeling was associated with gland cell endoplasmic reticulum, (-)-trans-isopiperitenol dehydrogenase labeling was restricted to secretory cell mitochondria, while (+)-pulegone reductase labeling occurred only in secretory cell cytoplasm. We discuss this pathway compartmentalization in relation to possible mechanisms for the intracellular movement of monoterpene metabolites, and for monoterpene secretion into the extracellular essential oil storage cavity.     

Comparative analysis of ultrastructure of glandular trichomes in two Nepeta cataria chemotypes (N. cataria and N. catena var. citriodora)

Glandular trichomes from the leaf surface of Nepeta cataria and N. cataria vai.citriodora have been examined using transmission and scanning electron microscopy. Peltate glands and capitate hairs type I were found on leaves of N. cataria. Both types had single stalk cells. Leaves of N. cataria var. citriodora bore peltate glands with unicellular or bicellular stalk, capitate hairs type I (with unicellular stalk) and capitate hairs type II (with unicellular or bicellular stalk). Peltate glands of N. cataria and of N. cataria var. citriodora were characterized by numerous leucoplasts sheathed by smooth reticular tubules and smooth endoplasmic reticulum; they are proposed to synthesize terpenes. The secretory cells of capitate hairs type I of N. cataria and those of N. cataria var. citriodora had well developed rough endoplasmic reticulum and dictyosomes. They had plastids with protein inclusions. These glands are supposed to produce slime. Capitate hairs type II of N. cataria var. citriodora had no analogs in N. cataria. Their secretory cells exhibited abundance of tubular endoplasmic reticulum and had unsheathed plastids with starch grains. Probably, these glands synthesize terpenes. The results of the study indicate that there is an obvious difference both in morphology and in ultrastructure of glandular trichomes in different chemotypes of N. cataria.     

Capitate glandular trichomes of Helianthus annuus (Asteraceae): ultrastructure and cytological development

Previous studies have shown that capitate glandular trichomes (CGT) of the common sunflower, Helianthus annuus, produce sesquiterpene lactones (STL) and flavonoids, which are sequestered and accumulated between the apical cuticle and the wall of the tip cells. To explore the cellular structures required and putatively involved in the STL biosynthesis and secretion, the present study was focused on the development of CGT and the comparison of the ultrastructure of its different cell types. Gradual maturation of flowers in the capitulum of the sunflower provided the possibility to study the simultaneous differentiation from the primordial to the secretory stage of CGT located by light microscopy (bright field, differential interference contrast and fluorescence) as well as transmission electron microscopy. It was shown that the CGT of sunflower anthers had a biseriate structure with up to 14 cell pairs. In mature trichomes, the apical cells called secretory cells were covered entirely by a large cuticle globe, which enclosed the resinous terpenoids and was specialised in thickness and structure. The secretory cells lacked chloroplasts and contained mainly smooth endoplasmic reticulum (sER). Conspicuous cell wall protuberances and an accumulation of mitochondria nearby occurred in the horizontally oriented cell walls. The cytological differences between stalk cells and secretory cells indicate a different function. The dominance of sER suggests its involvement in STL biosynthesis and cell wall protuberances enlarge the surface of the plasmamembrane of secretory cells and may be involved in the secretion processes of STL into the subcuticular space.     

Developmental and comparative ultrastructure of glandular hairs in the two Urticaceae. I. Urtica dioica

Secretion produced by glandular hairs is deposited mainly in the periplasmic space of the head cells. It stains intensely for both proteins and polysaccharides. The ultrastructure of meristematic, differentiating, mature and senescent head cells as well as the stalk and basal cells has been described in comparison to that in other cell types of the leaf. The specific features of the head cells are the proliferation of the granular endoplasmic reticulum as well as the multiplication of the dictyosomes and mitochondria during transition to the secretion stage. However, the frequency of dictyosomes varies among secreting hairs. The ER produces neither secretory nor transition vesicles and does not anastomose with the plasmalemma. In the absence of transition vesicles, the transport of secretory proteins and enzymes of polysaccharide synthesis from the ER to dictyosomes apparently includes the cytosolic step. Dictyosomes, though not appearing hypersecretory, produce two types of smooth secretory vesicles generated by the trans Golgi reticulum. The vectorial transfer of prosecretion and membranes across the dictyosome stack proceeds via the transport (shuttle) vesicles. It is, therefore, concluded that exocytosis of smooth secretory Golgi vesicles is the sole mechanism of release of both proteins and polysaccharides. Coated vesicles occasionally seen near the plasmalemma are likely to be involved in the endocytotic membrane retrieval. The secretion product disappears during senescence of the hairs and the secretory cells undergo vacuolation by means of local autophagy.     

Limonene Synthase, the Enzyme Responsible for Monoterpene Biosynthesis in Peppermint, Is Localized to Leucoplasts of Oil Gland Secretory Cells

Circumstantial evidence based on ultrastructural correlation, specific labeling, and subcellular fractionation studies indicates that at least the early steps of monoterpene biosynthesis occur in plastids. (4S)-Limonene synthase, which is responsible for the first dedicated step of monoterpene biosynthesis in mint species, appears to be translated as a preprotein bearing a long plastidial transit peptide. Immunogold labeling using polyclonal antibodies raised to the native enzyme demonstrated the specific localization of limonene synthase to the leucoplasts of peppermint (Mentha × piperita) oil gland secretory cells during the period of essential oil production. Labeling was shown to be absent from all other plastid types examined, including the basal and stalk cell plastids of the secretory phase glandular trichomes. Furthermore, in vitro translation of the preprotein and import experiments with isolated pea chloroplasts were consistent in demonstrating import of the nascent protein to the plastid stroma and proteolytic processing to the mature enzyme at this site. These experiments confirm that the leucoplastidome of the oil gland secretory cells is the exclusive location of limonene synthase, and almost certainly the preceding steps of monoterpene biosynthesis, in peppermint leaves. However, succeeding steps of monoterpene metabolism in mint appear to occur outside the leucoplasts of oil gland cells.     

ULTRASTRUCTURAL DEVELOPMENT OF CAPITATE GLANDULAR HAIRS OF CANNABIS SATIVA L. (CANNABACEAE)

The capitate-sessile and capitate-stalked glands of the glandular secretory system in Cannabis, which are interpreted as lipophilic type glandular hairs, were studied from floral bracts of pistillate plants. These glands develop a flattened multicellular disc of secretory cells, which with the extruded secretory product forms the gland head and the auxiliary cells which support the gland head. The secretory product accumulates beneath a sheath derived from separation of the outer wall surface of the cellular disc. The ultrastructure of secretory cells in pre-secretory stages is characterized by a dense ground plasm, transitory lipid bodies and fibrillar material, and well developed endoplasmic reticulum. Dictyosomes and dictyosome-derived secretory vesicles are present, but never abundant. Secretory stages of gland development are characterized by abundant mitochondria and leucoplasts and by a large vacuolar system. Production of the secretory product is associated with plastids which increase in number and structural complexity. The plastids develop a paracrystalline body which nearly fills the mature plastid. Material interpreted as a secretion appears at the surface of plastids, migrates, and accumulates along the cell surface adjoining the secretory cavity. Extrusion of the material into the secretory cavity occurs directly through the plasma membrane-cell wall barrier.     

线纹香茶菜叶上腺毛发育的细胞学研究

为进行中药溪黄草基原植物的品种鉴定,采用光镜和电镜对线纹香茶菜(原变种)[Isodon lophanthoides var.lophanthoides]叶上腺毛的发育进行细胞学研究。结果表明,线纹香茶菜具有头状腺毛和盾状腺毛2种类型。头状腺毛无色透明,由1个基细胞、1个柄细胞和1或2个头部分泌细胞构成;盾状腺毛为红色,由1或2个基细胞、1个柄细胞和4~8个分泌细胞构成头部。2种腺毛均由原表皮细胞经两次平周分裂形成,后因柄细胞和头部细胞所处的分化状态不同而形成两类腺毛。2种腺毛超微结构表明,质体、高尔基体和粗面内质网为主要分泌物产生和运输的细胞器。当盾状腺毛成熟时,角质层下间隙充满了分泌物,其分泌物的性质很可能决定了线纹香茶菜腺毛的颜色。     

Glandular trichomes of <Emphasis Type="Italic">Tussilago Farfara</Emphasis> (Senecioneae,Asteraceae)

Main conclusion

The glandular trichomes are developed on the aerial organs of Tussilago farfara ; they produce phenols and terpenoids. Smooth endoplasmic reticulum and leucoplasts are the main organelles of the trichome secretory cells. The aim of this study was to characterise the morphology, anatomy, histochemistry and ultrastructure of the trichomes in Tussilago farfara as well as to identify composition of the secretory products. Structure of trichomes located on the peduncles, bracts, phyllaries, and leaves were studied by light and electron microscopy. The capitate glandular trichomes consist of a multicellular head and a biseriate long stalk. Histochemical tests and fluorescence microscopy reveal phenols and terpenoids in the head cells. During secretory stage, the head cells contain smooth and rough endoplasmic reticulum, Golgi apparatus, diversiform leucoplasts with opaque contents in lamellae, chloroplasts, mitochondria, and microbodies. In the capitate glandular trichomes of T. farfara subcuticular cavity is absent, unlike glandular trichomes in other Asteraceae species. For the first time, content of metabolites in the different vegetative and reproductive organs as well as in the isolated capitate glandular trichomes was identified by GC–MS. Forty-five compounds, including organic acids, sugars, polyols, phenolics, and terpenoids were identified. It appeared that metabolite content in the methanol extracts from peduncles, bracts and phyllaries is biochemically analogous, and similar to the metabolites from leaves, in which photosynthesis happens. At the same time, the metabolites from trichome extracts essentially differ and refer to the above-mentioned secondary substances. The study has shown that the practical value of the aerial organs of coltsfoot is provided with flavonoids produced in the capitate glandular trichomes.

     

The Ultrastructure of Glandular Trichomes ofPhillyrea latifoliaL. (Oleaceae) Leaves

The anatomy and ultrastructure of glandular trichomes at differentdevelopmental stages were investigated inPhillyrea latifoliaL.leaves by transmission electron microscopy and histochemicaltechniques. The trichome consisted of a multicellular secretoryhead, a unicellular stalk and a collecting cell surrounded byepidermal cells and spongy mesophyll cells. There were numerousplasmodesmata across the cell walls of trichome cells, and especiallybetween the stalk cell and the collecting cell. The collectingcell and stalk cell contained few chloroplasts. Mitochondria,elements of the endoplasmic reticulum and small vacuoles wereabundant in the secretory cells. Crystals were present in thesecretory cells and the collecting cell, especially at the matureand senescent stages of trichome development. As the cuticle,which covered the secretory cells, did not show pores or perforations,it is proposed that secretion occurred by accumulation of productsin subcuticular spaces followed by diffusion through the cuticle.Callose accumulation was observed between the stalk cell andthe collecting cell of senescent trichomes, especially in salt-treatedplants. Trichome ontogeny was accelerated in salt-treated plants.Copyright1998 Annals of Botany Company Cuticle;Phillyrea latifolia; secretion; transmission electron microscopy; trichome development.     

Development of trichomes in the Abutilon nectary gland

Nectary trichomes of Abutilon striatum var. thompsonii arise by sequential periclinal divisions of outpushings from epidermal cells so producing trichomes that, when mature, are about 12 cells long. All epidermal cells within the nectary undergo this transformation. Later, anticlinal divisions lead to a multiseriate lower part of the trichome. The original epidermal cell becomes the basal cell which increases substantially in volume during development, thus leading to lateral separation of the trichomes. Above the basal cell is the stalk cell which develops an apoplastic barrier in its anticlinal (outer) wall. Secretion ultimately takes place from a capitate tip cell. An initially very thin cuticular layer, which overlies the whole trichome, eventually becomes as thick as the cell wall itself (approx. 0.4 μm). The pre-secretory hairs contain numerous small, condensed mitochondria; poorly differentiated plastids; dictyosomes with coated vesicles; small vacuoles; and a large amount of smooth endoplasmic reticulum ("secretory reticulum") which contrasts with the rough endoplasmic reticulum seen during earlier developmental stages. As secretion proceeds, vacuolation becomes more extensive. Plasmodesmata are present between all the cells of the trichome and diminish in frequency from about 12.0 μm^-2 in the stalk cell to about 4.0 μm^-2 in the apical cells. This variation in plasmodesmatal frequency along the trichome is seen at all stages of development. The ultrastructural evidence would be consistent with the hypothesis that the pre-nectar flows through the plasmodesmata from cell to cell, is loaded into a "secretory compartment", and is then unloaded into the apoplast from all cells of the trichome distal to the stalk cell.     

ULTRASTRUCTURE OF GLANDULAR OVARIAN TRICHOMES OF CYPRIPEDIUM CALCEOLUS AND C. REGINAE (ORCHIDACEAE)

Trichomes on the orchid ovary are a possible site of synthesis and secretion of the floral scent. Scanning electron microscopy of these trichomes shows a bulbous cell on a two-celled stalk. Thin sections of the tip cell revealed the morphology of an active, secretory cell with unusual coated vesicles in the extra-cellular deposition. Abundant smooth endoplasmic reticulum (ER) aggregated beneath the plasma membrane in the apical region of the cell and the limited dictyosomes in the cell suggest direct secretion by ER. Numerous lipid droplets are present in the apical area. Plastids, found only in the basal region of this cell, are more round in profile than typical chloroplasts and contain only a few unstacked thylakoids and a limited membranous reticulum. In addition to the normal plastid envelope, a double layer of membrane (probably ER) is tightly appressed to each dense, starch-free plastid. Highly specialized morphology and subcellular localization of organelles suggest the secretory nature of these trichomes.     

Ultrastructural Aspects of the Nectary Spur of Limodorum abortivum (L) Sw. (Orchidaceae)

The ultrastructure of the nectary spur of Limodorum abortivum(L) Sw. was examined before and after anthesis. In cross sectionthe nectary spur shows an internal epidermal layer of thin-walledcells bordering the secretory cavity and 10–12 layersof parenchyma cells. The ultrastructure of the secretory cellssuggests the involvement of ER, Golgi and plastids in nectarsecretion. The nectar accumulated in the sub-cuticular spaceis released into the nectariferous cavity by rupture of theouter layer of the cuticle. Limodorum abortivum (L) Sw., Orchidaceae, nectary spur, nectar secretion, ultrastructure, anthesis, endoplasmic reticulum, dictyosomes, plastids     

Leaf glandular trichomes in Empetrum nigrum: morphology,histochemistry, ultrastructure and secondary metabolites

The morphology, histochemistry and ultrastructure of the glandular trichomes on Empetrumnigrum leaves have been studied and more than a third of the metabolites were identified. Samples of the leaves were fixed and processed for light and electron microscopy. Glandular trichomes are situated on the inner surface of the rolled leaves. They have a clavate head and a short stalk. Histochemical tests and fluorescent microscopy demonstrate differentiated staining of the various cell types in the glandular trichome for proteins, pectins, lipids, tannins and phenylpropanoids. During secretion, the secretory cells contain rough and smooth endoplasmic reticulum, Golgi stacks with large vesicles, diversiform leucoplasts in contact with a reticular sheath and opaque deposits in the vacuoles. There are ultrastructural and functional differences between the secretory cells in the trichome head: synthesis of hydrophilic substances predominately occurs in the upper and middle secretory cells, whereas synthesis of lipophilic compounds takes place in the middle and lower secretory cells. Gas chromatography–mass spectrometry was used to determine the content of metabolites in the methanol extracts from the leaves. Many phenolic compounds (phenolic acids, bibenzyls, catechins, flavanones and flavan‐3‐ols) as well as several terpenoids were found. Two chalcones (2′,4′‐dimethoxydihydrochalcone and 2′,4′,6′‐trihydroxydihydrochalcone), one bibenzyl (batatasin III), one flavanone (7‐hydroxyflavanone) and 8 terpenoids (including phytol, α‐tocopherol, ß‐sitosterol, α‐amyrin, uvaol, oleanolic acid, ursolic acid and dehydroursolic acid) were identified in E. nigrum leaf extracts. The total yield of phenolic compounds is five to six times higher than the yield of terpenoids. It has been established that chalcones have no hydroxyl groups in ring B whereas bibenzyls have a hydroxyl group in the 3‐position in ring B. On the basis of the histochemistry, fluorescent microscopy, ultrastructure and chemical analysis, it may be concluded that synthesis and accumulation of phenolic substances and terpenoids takes place in the clavate glandular trichomes. Secondary metabolites synthesized in the trichomes protect leaf tissues from viruses, bacteria and pathogenic fungi.     

Development of secretory activity in the seminal vesicle of the male migratory grasshopper,Melanoplus sanguinipes (fabr.) (Orthoptera : Acrididae)

This paper describes the ultrastructure of the seminal vesicle and the isoelectric focusing patterns of its secretion during sexual maturation and after allatectomy in Melanoplus sanguinipes (Fabr.) (Orthoptera : Acrididae). In epithelia from seminal vesicles of newly fledged males, the rough endoplasmic reticulum is well developed, and Golgi complexes are elaborate, which indicates the gland is metabolically active. The cells also contain large glycogen deposits and the lumen microvilli are well differentiated. These ultrastructural features are more dominant in 24-hr-old adults where the cytoplasm is clearly differentiated into basal and apical regions. Basally, the cytoplasm is dominated by rough endoplasmic reticulum, large Golgi complexes, glycogen deposits and numerous mitochondria, while the apical cytoplasm is filled with large secretory and/or lysosomal vesicles. Between days 3 and 7, the ultrastructural features change little other than the rough endoplasmic reticulum cisternae, which become vesicular. Analysis by isoelectric focusing shows that the amount of secretory protein increases with age until day 3, at which time the gland contains its full complement of secretion. In seminal vesicles from allatectomized insects, ultrastructural features of cells and isoelectric focusing patterns of the secretion arc identical to those from normal males.     

Light and electron microscopy study of the salivary glands of the carnivorous opisthobranch Philinopsis depicta (Mollusca, Gastropoda)

Cephalaspideans are a group of opisthobranch gastropods that comprises carnivorous and herbivorous species, allowing an investigation of the relationship between these diets and the morphofunctional features of the salivary glands. In this study, the salivary glands of the carnivorous cephalaspidean Philinopsis depicta were observed by light and electron microscopy. The secretory epithelium of these ribbon-shaped glands is formed by ciliated cells, granular cells and cells with apical vacuole. In ciliated cells the nucleus and most cytoplasmic organelles are located in the wider apical region and a very thin stalk reaches the base of the epithelium. These cells possess significant amounts of glycogen. Granular cells are packed with electron-dense secretory granules and also contain several cisternae of rough endoplasmic reticulum and Golgi stacks. The other type of secretory cell is mainly characterized by the presence of a large apical vacuole containing secretion. These cells possess high amounts of rough endoplasmic reticulum cisternae and several Golgi stacks. Vesicles with peripheral electron-dense material are also abundant, and seem to fuse to form the apical vacuole. The available data point out to a significant difference between the salivary glands of carnivorous and herbivorous cephalaspidean opisthobranchs, with an intensification of protein secretion in carnivorous species.     

羽叶薰衣草表皮毛的发育解剖学研究

对羽叶薰衣草(LavandulapinnataL.)茎和叶上两种表皮毛(腺毛和非腺毛)发育的解剖学观察表明,两者的发生都源于茎或叶的原表皮细胞,但外部形态、发育过程及功能明显不同。腺毛有头状腺毛和盾状腺毛两种类型,均由1个基细胞、1个柄细胞和头部细胞构成。头状腺毛的头部只有1个或2个分泌细胞,盾状腺毛由8个分泌细胞构成头部。非腺毛由3-20个细胞组成,可分为三种类型:单列不分枝、二叉分枝和三叉及三叉以上多分枝的树状分枝。非腺毛的顶部细胞由基部到顶部逐渐变细,先端成尖形。腺毛发育由原表皮细胞经两次平周分裂形成,由于柄细胞和头部细胞所处的分化状态不同而发育成两类腺毛。非腺毛由非腺毛原始细胞经二次或多次平周分裂和不均等分裂,再发育成数个至二十多个子细胞。     

Ultrastructure of the sexual segments of the kidney in male and female lizards,Cnemidophorus l. lemniscatus (L.)

Summary Kidneys of adult male and female lizards were studied by electron microscopy, in order to understand the ultrastructure of the collecting duct and a differentiated part thereof, the sexual segment, which is an important accessory sexual organ. First portion of sexual segment in males: The cells are filled with large secretory granules of a wide range of opacities. The granular endoplasmic reticulum is abundant; basal formations of superimposed flat cisternae are frequent. Distended vesicles and microvesicles prevail in the supranuclear, well developed Golgi apparatus. Evidences indicate that secretion of these cells is holocrine. Second portion of sexual segment in males: All of the secretory granules are apical in location and relatively electron-opaque; they show a denser core. This core is formed by a substance which, after lying in contact with ribosomes, enters the secretory vesicles of the highly developed Golgi apparatus. A lighter substance is then condensed around it. The secretion of the granules is merocrine. The granular endoplasmic reticulum is very abundant in these cells, but basal ergastoplasmic formations are lacking. Sexual segment in females: The cells show features similar to those of the male first portion, but they are smaller. Undifferentiated collecting duct: Most of the cells are mucigenic. They have small ovoid, apical secretory granules. The density of the granules varies from cell to cell; when they are electron-lucent, they exhibit laminar or dotted opaque figures. Moderately developed Golgi apparatus and granular endoplasmic reticulum, as well as elongated mitochondria, occur in mucigenic cells. Intercalated among the latter are non-secretory cells. They have very abundant mitochondria, numerous microvilli, many pinocytic and smooth-membrane vesicles, whereas the organelles participating in synthetic processes are poorly developed; their function is most likely related to active solute transport.     

Anatomical studies of the secretory structures: Glandular trichomes and ducts, in Grindelia pulchella Dunal (Astereae, Asteraceae)

The ultrastructure of the glandular trichomes and secretory ducts of Grindelia pulchella was studied. Plastids, mitochondria and endoplasmic reticulum are involved in the secretory process of both, trichomes and ducts. A special tissue with “transfer cells” is associated with the duct epithelial cells. The secretion is produced in the transfer cells and then is transferred to the duct epithelial cells where it accumulates in the vacuoles. The occurrence of cavities within the cell walls of the trichome cells and duct epithelial cells is described. The secretion is accumulated between the cell wall and the cuticle of these cells. When the cuticle is broken the secretion is released. We conclude that granulocrine secretion operates in this species.