Apocrine secretion of amylase and exocytosis of trypsin along the midgut of Tenebrio molitor larvae

Amylase and trypsin were purified from Tenebrio molitor midgut larvae and used to raise antibodies in a rabbit. A Western blot of T. molitor midgut homogenates, after sodium dodecyl sulfate-polyacrylamide gel electrophoresis using amylase and trypsin antisera, showed only bands co-migrating with the purified enzymes. The antisera were used to localize the enzymes by immunofluorescence and immunogold labeling. Amylase occurs in a few regularly disposed anterior midgut cells. Non-amylase-secreting anterior midgut cells are proposed to be water-absorbing cells based on morphology and dye movements. Amylase is found inside vesicles originating from Golgi areas that seem to fuse together before their release along with the now disorganized apical cytoplasm (apocrine secretion). Trypsin precursors are observed inside small vesicles near the apical plasma membrane of posterior midgut cells, suggesting an exocytic mechanism of secretion, followed by putative trypsin activation. Apocrine secretion is thought to be an adaptation to enhance the dispersion of secretory vesicle contents released from a water-absorbing epithelium, whereas exocytosis is an efficient secretory mechanism in a water-secreting epithelium.     

Ultrastructural changes in the oviductal mucosa throughout the sexual cycle in Bufo arenarum

In Bufo arenarum the oviduct exhibits conspicuous changes throughout the sexual cycle. In the present study, we analyzed the optical and ultrastructural characteristics of the oviductal pars convoluta mucosa, the portion responsible for jelly secretion, during both the preovulatory and postovulatory periods. Secretory epithelial cells, ciliated cells, basal cells, and glandular cells are described. Secretory epithelial cells are characterized by the presence of secretory granules, the size, shape and electron density of which vary markedly. Their contents are mainly released by exocytosis into the oviductal lumen. Moreover, in the preovulatory period, apocrine, and holocrine secretion processes frequently occur. During the postovulatory period, these cells exhibit a marked diminution of secretory granules. Ciliated cells show a typical ultrastructural organization. Basal cells are distinguished in the lower part of the epithelium by their heterochromatic nuclei and electron‐lucent cytoplasm. These cells, to the best of our knowledge, are reported for the first time in Amphibia. Glandular cells exhibit oval, round, or polyhedric granules, most of them with one or more cores. Our results indicate that the contents of epithelial and glandular secretory cells are partially secreted during the preovulatory period. Additional secretion occurs during the transit of the oocytes. J. Morphol. 239:61–73, 1999. © 1999 Wiley‐Liss, Inc.     

小地老虎雄性附腺细微结构和功能及高温的影响

本文通过光镜、电镜和生化分析等方法,研究了小地老虎Agrotis ypsilon(Rottemberg)雄性附腺的细微结构和功能,结果表明：(1)雄性附腺是一对管状腺,基段粉红色,中段桔红色,端段乳白色。形态分化在蛹前期完成,分泌功能在羽化后4天进入旺盛期;(2)附腺细胞属蛋白质合成型,具有旺盛的合成蛋白质的能力,胞内含有致密的粗面内质网和游离核糖体颗粒,大量的分泌液泡均匀地分布在细胞质中;(3)顶浆分泌和局部分泌是腺体的二种主要分泌方式, 前者分泌的颗粒物为糖蛋白性质(Pas阳性),后者吩泌的网状物为非糖蛋白性质(Pas阴性),二者在腺腔内呈有规则的放射状排列“4”雄性附腺分泌物具有种的特异性,小地老虎、棉铃虫和粘虫等夜蛾科昆虫分泌物的蛋白电泳谱带存在明显的种间差异,高温(32℃)抑制了雄性附腺分泌某些蛋白质的能力,从而改变精液的成分。     

麝泌香盛期后麝香腺囊的显微与超微结构和麝香分泌研究

麝香是雄麝特有的麝香腺囊分泌物,是对麝群行使化学通讯机能的一种信息素。本文研究了泌香盛期后麝香腺囊的显微与超微结构,探索了雄麝年周期的麝香分泌及形成机理。     

Secretion mode of the harderian gland of rats after stimulation by cholinergic secretagogues.

We studied the morphological changes in rat Harderian glands 30 min after injection of cholinergic secretagogues. In controls, the glands exhibited a tubuloalveolar structure with relatively wide lumina, in which some osmiophilic dense droplets exocytosed from glandular cells were observed. Also two types of glandular cells (type A cells and type B cells sometimes showing exocytotic figures of lipid-secretory vacuoles) and myoepithelial cells were recognized. After injection of carbamylcholine chloride (subcutaneously, 0.1 mg/kg body weight), which has both nicotinic and muscarinic actions, many of the alveolar lumina dilated and contained a small number of osmiophilic droplets. Exocytotic figures in both types of cells and a pronounced decrease in the number of vacuoles in the glandular cells were observed. However, there was no evidence of apocrine or holocrine secretion. The injection of the higher dose of carbamylcholine (1.0 mg/kg) caused fusion of secretory vacuoles in the apical cytoplasm and contraction of myoepithelial cells. Most alveoli showed no clear lumina; their centers were jammed with cytoplasmic fragments and accumulated secretory products. Massive discharge of cytoplasmic fragments containing some secretory vacuoles was often observed. This may be classified as apocrine secretion. Bethanechol chloride (subcutaneous injection, 1.0 mg/kg), a muscarinic agonist, stimulated the Harderian-gland secretion, and enhanced exocytosis was observed. The discharge from the glandular cells, following injection of various doses of carbamylcholine, were almost inhibited by atropine sulfate, a muscarinic antagonist. The present results suggested that the cholinergic systems regulate the secretion of rat Harderian-gland cells which have muscarinic receptors.     

Cutaneous glands of male and female impalas (Aepyceros melampus): seasonal activity changes and secretory mechanisms

The cutaneous glands of the forehead and the metatarsus were studied by histological and histochemical methods and electron microscopy in adult male and female impalas in various seasons of the year. All glandular areas consist of apocrine and holocrine glands, which, however, occur in different proportions. Our findings in the apocrine gland cells suggest (1) the synthesis and exocytosis of a glycoproteinaceous secretory product stored in secretory granules, (2) typical apocrine secretion of the transformed apical cytoplasm, and (3) transepithelial fluid transport. The Golgi apparatus and apical membrane have binding sites for several lectins (PNA, HPA, RCA I, WGA). Cytokeratins 7, 14 and 19 are expressed at various intracellular localizations, suggesting an active role in the secretory mechanisms. The glands of the male forehead show marked seasonal changes in activity that are correlated with the main phases of the reproductive cycle, with the highest cellular activity occurring during the rut in April/May. The female forehead glands are only moderately developed and do not undergo seasonal changes. The metatarsal glands are of equal size in males and females and show no seasonal changes in activity. This study supports the hypothesis that (1) forehead glands in the male have a signaling role in the rut and (2) the metatarsal glands have a more general, probably social role maintaining and restoring contact between herd members.     

Mitochondrial deformation and apocrine secretory mechanism in the rabbit submandibular organ as revealed by electron microscopy

Summary The submandibular organ (a sort of apocrine sweat glands) of the rabbit was observed with the electron microscope. The cell structure of glandular tubules varies depending upon the secretory activity; there are three functional stages. The secretory cells at the resting stage are characterized by low height, absence of secretory substance, and presence of small and slender mitochondria.In the synthesizing stage, enlargement and peculiar deformation of mitochondria are observed. Secretory substance always occurs near the deformed mitochondria. The part of a mitochondrion closely abutting on the secretion mass is extremely thin, and contains longitudinally oriented cristae. Sometimes a direct continuity is observed between the thinned portion of the deformed mitochondria and the mass of secretory substance. It is presumed that the secretion is initially produced in the mitochondria and then discharged from them. The Golgi apparatus and the rough surfaced endoplasmic reticulum may be involved indirectly. Smooth surfaced vesicles, probably related to the transport of raw material, are extremely abundant in the cells of this stage.The development of a generally homogeneous projection into the gland lumen is characteristic of the stage of secretion discharge. The mitochondria are again small and slender, and the secretion is liquefied. At the base of the full-grown projection, cytoplasm is condensed to form a demarcation zone from which the projection may become detached. This mechanism of release of secretory product is quite the same as the so-called apocrine secretory process long postulated by light microscopists.     

Fine structure of the axillary gland in the brown bullhead (Ictalurus nebulosus)

The axillary glands of Ictalurus are lobulated invaginations of the epidermis, opening at a pore between the pectoral spine and the cleithrum. Holocrine cells lining a false lumen form a viscous secretion. The secretory cells originate in the tenuous basal layer of the gland wall. Secretion is initiated by the formation of compound vesicles in cells that become very large and have complex cytoplasm of a varied appearance. Golgi systems are well developed and the perinuclear cytoplasm may contain many mitochondria and sacs of ribosomal endoplasmic reticulum; some tracts of cytoplasm are vesicular and contain free ribosomes. Some cells contain numerous large lysosomes, and some have extensive contents of fibrillar masses imperfectly separated by membranes, that recall the appearance of the mucous secretion of goblet cells. The secretory cells break down, releasing the degenerating organelles, including the nuclei, into the false lumen. Some structures are still recognizable in the secretion even after it has been expelled, but the main part of the formed secretion consists of the mucus-like masses. Various leucocytes are found in the gland walls and embedded in the secretion. The fine structure differentiates the holocrine cells of the axillary gland from the club cells of the epidermis, and from the venom glands associated with the fin spines of catfishes. The function of the axillary gland secretion remains unknown.     

Acid phosphatase localization in the digestive glands of Dionaea muscipula Ellis flytraps

The intracellular localization of acid phosphatases in stimulated digestive glands of Dionaea flytraps has been studied to provide evidence for the route taken by this enzyme during secretion. Previous studies have either included or excluded a role for the dictyosomes in this pathway. Both p-nitrophenyl phosphate and beta-glycerophosphate were used as substrates, and both gave similar localization patterns. Unstimulated glands contained little phosphatase activity in the endomembrane system, whereas 24 and 48 hr after stimulation, heavy deposits of lead were located in the endoplasmic reticulum cisternae, including the nuclear envelope, the dictyosome cisternae, and secretory vesicles. Since dictyosome activation, as judged by the presence of secretory vesicles in the cytoplasm, also coincides with gland stimulation, we conclude that secretion of the hydrolase enzymes occurs via this route and not, as suggested elsewhere, via direct endoplasmic reticulum to plasma membrane connections.     

Microstructural Analysis of the Capture Thread Spinning Apparatus in Orb Web Spiders

The microstructural characteristics of the capture thread production from silk glands in the orb web spiders were analyzed using scanning and transmission electron microscopes. Sticky and gluey capture threads of the web are originated from the silks of two flagelliform glands and four aggregate glands. They supply precursors of the secretory silks to a pair of characteristic “triad” spinning units on the posterior spinnerets. The aggregate gland is composed of large and multi‐lobed secretory region and thick excretory duct surrounded by large irregular nodules. The excretory duct of this gland basically consists of three superposed types of cells which are inner columnar epithelium, nodule forming cells and outer connectives. The nodules contain numerous mitochondria and glycogen particles within their cytoplasm and they are surrounded by the same sheath of thin connective tissues. Secretory region of the aggregate gland which produce water‐soluble components of the capture thread comprises discrete secretory vesicles and extensive rough endoplasmic reticulum. Characteristically, secretory droplets are formed without involvement of the Golgi complexes, suggesting that they do not play an important role in the processing of the capture threads. However the electron densities and internal textures of the granules are observed with diverse according to their maturation level. Finally, the secretory products are released by the mechanism of apocrine secretion losing part of their cytoplasm during this process.     

Ultrastructural changes in the oviductal epithelium of Merino ewes during the estrous cycle

Isthmic and ampullary oviductal epithelia sampled from Merino ewes at days -1, 1, 3, and 10 of the estrous cycle (estrus = day 0) were studied by scanning and transmission electron microscopy after fixation by vascular perfusion. Secretory cells, ciliated cells, and lymphocytelike basal cells were observed in both isthmic and ampullary epithelium at all stages of the estrous cycle studied and their ultrastructural features were analyzed. Synthesis of lamellated secretory granules occurred in the ampullary secretory cells during the follicular and early luteal phases, and their contents were released by exocytosis into the oviductal lumen during the luteal phase. Granule release was associated with nucleated apical protrusion of these cells into the oviductal lumen. No such secretory activity was displayed by isthmic secretory cells even though a few cells contained nonlamellated granules. Apocrine release of apical vesicles and accompanying cytoplasmic material from apical protrusions of ciliated cells occurred in the isthmus around estrus but not in the ampulla. This unexpected feature has not previously been reported in any other mammal. Dendritic basal cells were distinguished in the lower part of the epithelium by their heterochromatic nuclei, electron-lucent cytoplasm, and lack of attachment zones. No migration of basal cells was observed, and their ultrastructural features were similar in the ampulla and isthmus and at all stages of the estrous cycle examined. The function of these lymphocytelike cells in the epithelium is uncertain, but the presence of phagocytic bodies and lysosomes in 20% of them may indicate a phagocytic role.     

Human axillary apocrine glands: proteins involved in the apocrine secretory mechanism

The apocrine secretory mechanism is a mode of secretion by which the apical part of the cell cytoplasm is pinched off, which leads to the formation of an aposome. The distinct mechanism of formation and decapitation of the aposome is not well investigated. Only few proteins are known that are involved in this secretory mechanism. We studied the human axillary apocrine gland and looked at proteins associated with cytokinesis, a process that is comparable to the pinching-off mechanism of apocrine glandular cells. By immunohistochemistry, we detected actin, myosin II, cytokeratin 7 and 19, α- and β-tubulin, anillin, cofilin, syntaxin 2, vamp8/endobrevin and septin 2. In highly active glandular cells, these proteins are located at the base of the apical protrusion when the aposome is in the process of being released or are concentrated in the cap of the apical protrusion. These findings demonstrate new insights on apocrine secretory mechanisms and point to similarities to the terminal step of cytokinesis, which is regulated by a SNARE-mediated membrane fusion event.     

Ultrastructural and histochemical observations on secretory phenomena in the resting human mammary gland

Ultrastructure and histochemistry of clinically normal appearing tissue and secretions of non-lactating human mammary glands have been investigated in order to document and analyse secretory phenomena in the resting gland. The material studied originated from women of different ages (18-74 years) who underwent plastic surgery or surgery for various disorders of the breast. The epithelia of small ducts and of alveolar enlargements as well as acini contained moderate amounts of mitochondria and of cisterns of the rough endoplasmic reticulum; the transcisterns of the Golgi apparatus which were surrounded by smooth and coated vesicles, exhibited modest dilatations, the number of lysosomes increased with age; regularly glycogen particles and bundles of intracellular filaments (phi 5 nm) were to be observed. Typical casein vesicles and stages of apocrine secretion of milk fat globules were not seen. The following features indicated secretory activity: differently sized vesicles and granules with flocculent, dense, or light contents were regularly to be seen in the apical cytoplasm often immediately below the apical plasma membrane of the epithelia of the small ducts and even more frequently in the alveolar enlargements. Secretory products of fine granular or filamentous structure, probably containing proteins, were frequently found within the lumen. Different types of lipid and liposome-like particles were detected both in intracellular localization as well as in ductal lumina. As demonstrated by lectin histochemistry the secretory products also contained a considerable amount of carbohydrate components. The composition of the secretory products of the resting gland is of clinical interest since the chronical deposition of secretions, which among others possibly contain enzymes producing oxygen radicals, may lead to pathological changes of mammary gland tissue.     

The ultrastructural investigation of the midgut in the quill mite Syringophilopsis fringilla (Acari,Trombidiformes: Syringophilidae)

The midgut of the females of Syringophilopsis fringilla (Fritsch) composed of anterior midgut and excretory organ (=posterior midgut) was investigated by means of light and transmission electron microscopy. The anterior midgut includes the ventriculus and two pairs of midgut caeca. These organs are lined by a similar epithelium except for the region adjacent to the coxal glands. Four cell subtypes were distinguished in the epithelium of the anterior midgut. All of them evidently represent physiological states of a single cell type. The digestive cells are most abundant. These cells are rich in rough endoplasmic reticulum and participate both in secretion and intracellular digestion. They form macropinocytotic vesicles in the apical region and a lot of secondary lysosomes in the central cytoplasm. After accumulating various residual bodies and spherites, the digestive cells transform into the excretory cells. The latter can be either extruded into the gut lumen or bud off their apical region and enter a new digestive cycle. The secretory cells were not found in all specimens examined. They are characterized by the presence of dense membrane-bounded granules, 2–4 μm in diameter, as well as by an extensive rough endoplasmic reticulum and Golgi bodies. The ventricular wall adjacent to the coxal glands demonstrates features of transporting epithelia. The cells are characterized by irregularly branched apical processes and a high concentration of mitochondria. The main function of the excretory organ (posterior midgut) is the elimination of nitrogenous waste. Formation of guanine-containing granules in the cytoplasm of the epithelial cells was shown to be associated with Golgi activity. The excretory granules are released into the gut lumen by means of eccrine or apocrine secretion. Evacuation of the fecal masses occurs periodically. Mitotic figures have been observed occasionally in the epithelial cells of the anterior midgut.     

Quantitative evidence for merocrine secretion in insect midgut cells

The occurrence of apical extrusions from cells has been observed on a large number of occasions in a wide variety of histological investigations. But whether the extrusions observed are a normal physiological event or are artefacts of the experimental procedures adopted has been the subject of much controversy. Lehane (1976a) described the extrusion of apical portions of the opaque zone midgut cells of Stomoxys calcitrans which contained large numbers of storage granules (merocrine secretion). This process involved the cell in the loss of cytoplasm, mitochondria and membranes as well as secretory products. If merocrine secretion is a normal physiological event rather than a fixation artefact then large falls should occur in the quantities of these components in the cell following the secretion burst. Using stereological methods the opaque zone cells were quantitatively described before and after the period in which merocrine secretion occurs (using these timings any possibility of producing apical extrusions as fixation artefacts was avoided). The results are entirely consistent with merocrine secretion having occurred. The volume of the cytoplasm falls by 23.74%, mitochondria by 30.55% and the total area of membrane in the cell by 32.59%.     

Serous gland polymorphism in the skin of<Emphasis Type="Italic"> Phyllomedusa hypochondrialis azurea</Emphasis> (Anura,Hylidae): response by different gland types to norepinephrine stimulation

Stimulation by norepinephrine in physiological concentration was used on the dorsal skin of the Argentine tree-frog Phyllomedusa hypochondrialis azurea to trigger contraction of myoepithelial cells encircling the serous glands and provoke secretory release. This hylid species possesses two kinds of serous cutaneous glands, producing secretory granules or vesicles (type Ia and Ib serous units, respectively), along with serous-derived glands which synthesise lipids and store them in complex aggregates (type II units). Structural and ultrastructural observations on myoepithelia, secretory units and gland products collected in saline after discharge, revealed consistent but different responses in the three types investigated. Type Ia glands reacted intensely to treatment, with both contractile and secretory responses, type Ib glands were only mildly affected in their myoepithelia and glands of type II were not affected at all. According to data available in the literature, these findings suggest that: (a) the dense (type Ia) granules are expelled as a phasic response through bulk (holocrine) discharge, (b) the secretory (type Ib) vesicles are released as a tonic response through a merocrine mechanism and (c) lipid (type II) aggregates are exuded as a secretory component of a complex behavioural response which tends to reduce transcutaneous water loss. Furthermore, these findings indicate that the use of pharmacological modulation of myoepithelial activity allows selective collection of skin products in species characterised by serous gland polymorphism.     

Sécrétions apocrines dans le tractus génital mâle: Roles potentiels dans la maturation des gamètes

Apocrine secretory vesicles have been observed in the lumen of several ducts of the male genital tract, namely the epididymis and vas deferens. Initially considered to be tissue-fixation artefacts, they are now recognized as genuine elements, which might play physiological roles in terms of sperm maturation. The aim of this review is to present these vesicles observed in various mammalian species and to describe their respective organization, content and putative roles.     

Nature's ingenuity: bypassing the classical secretory route via apocrine secretion

Although it has been suggested that epithelial cells of the male reproductive system are involved in apocrine secretion, this method of secretion is not fully understood. In the present study, apocrine secretion was investigated in epithelial principal cells lining the epididymis and vas deferens (VD) of adult mice. The tissues were fixed by cardiac vascular perfusion with glutaraldehyde for routine electron microscope (EM) analysis and Bouin's fixative for light microscope (LM) immunocytochemistry to access functional roles. In the epididymis and VD, the apex of principal cells revealed protrusions of cytoplasm referred to as apical blebs (ABs). The latter contained solely numerous free ribosomes, 20 nm vesicles and few ER cisternae, suggesting segregation of their contents. While some ABs displayed wide areas of contact with the apical principal cell cytoplasm, others showed thin stalk-like attachment points as well as fissures at the junction of the two areas. Together with images of ABs and their contents deep in the lumen, it is suggested that ABs detach from principal cells whereupon they breakdown to release their contents therein. As ABs of the epididymis were immunoreactive for glutathione-S-transferases (GSTs) and ubiquitin, it is proposed that these proteins are synthesized on free ribosomes in ABs and that apocrine secretion represents the manner whereby they enter the lumen to effectively protect sperm from free radical injury and ubiquitinate proteins for degradation, respectively. ABs of the VD were immunoreactive for 3beta-HSD, suggesting that they are also capable of synthesis of steroids with their release via apocrine secretion. Taken together the data provide evidence for apocrine secretion in the adult mouse epididymis and VD that could play important roles in relation to sperm maturation, protection and viability.     

Protein discharge from immature secretory granules displays both regulated and constitutive characteristics.

At physiological glucose concentrations, isolated pancreatic islets release a minor portion of their newly synthesized insulin and precursors in a phase of secretion which is largely complete by 4 h of chase. Discharge during this period can be amplified by secretagogues, yet is not abolished by conditions which fully suppress regulated release from dense core secretory granules. The ability to stimulate the secretion and the biochemical profile of released proinsulin-related peptides indicate that secretion during this period originates from immature granules. The stoichiometry of release of labeled C-peptide:insulin during this phase is 1:1 at high glucose concentrations. However, at physiologic or low concentrations, C-peptide is released in molar excess of insulin as if the exocytotic vesicles carrying this secretion were budding from a post-Golgi compartment in which the lumen was composed of condensing insulin and soluble C-peptide. These findings can be explained by a model for regulated secretory protein traffic in which direct exocytosis of young granules is stimulated by higher glucose concentrations and vesicle budding from immature granules occurs at lower concentrations. Thus, insulin targeting from immature granules exhibits both regulated and constitutive-like characteristics.