Intracellular pathway and kinetics of protein secretion in the coagulating gland of the mouse

The coagulating gland of male rodents is part of the prostatic complex. Various mechanisms of secretion have been postulated, in part because organelles commonly involved in the secretory process possess unusual features, such as extreme distension of the rough endoplasmic reticulum. In the present study, the pathway, kinetics, and mode of secretion in the coagulating gland of the mouse were studied by electron microscope autoradiography at intervals between 5 min and 8 h after administration of 3H-threonine. The percentage of grains associated with the rough endoplasmic reticulum was initially high and generally decreased throughout the experiment, while a pronounced rise in the proportion of grains associated with the Golgi apparatus and secretory granules was observed 6 h after injection of precursor. In addition, there was a smaller elevation in the percentage of grains over the Golgi apparatus and secretory granules between 1 and 4 h, and radioactive material first reached the lumen of the gland 4 h after injection of the precursor. Although the general pathway of intracellular transport of secretory protein resembles that in other cells, the results indicate that there are several unusual aspects to the secretory process in the coagulating gland. First, the rate of transport was markedly slower than in most other exocrine gland cells, since the bulk of the labeled protein did not reach the Golgi apparatus and secretory granules until 6 h after administration of precursor. This reflected prolonged retention of secretory products in the endoplasmic reticulum. Second, in addition to the major bolus of labeled material that traversed the cells at about 6 h, a smaller wave of radioactivity appeared to pass through the Golgi apparatus and secretory granules and reach the lumen earlier, within the first few hours after the injection. Finally, the primary mode of secretion in the coagulating gland appears to be merocrine because the secretory granules contained much labeled protein.     

Simultaneous apocrine and merocrine secretion in the rat coagulating gland

The coagulating gland of the rat synthesizes two prevalent secretory proteins (transglutaminase and 115 K) that are discharched in a different manner, one being secreted in an apocrine fashion (transglutaminase) and the other one in a merocrine way (115 K). Differences in the intra- cellular pathway and the release of either protein were studied using immunofluorescence on semithin sections, immunoelectron microscopy of preembedding-processed chopper sections and postembedding-processed ultrathin sections of rat coagulating gland. Immunohistochemical staining using an anti-transglutaminase antibody resulted in dense labeling of the cytoplasm of secretory cells and their apical blebs, whereas the cisternae of the rough endoplasmic reticulum and the Golgi apparatus were completely unlabeled. When, on the contrary, the anti-115 K antiserum was used, dense labeling of the cisternae of the rough endoplasmic reticulum, the Golgi apparatus, and the secretory granules was seen. Intraluminal secretion was also labeled, but the secretory blebs remained unlabeled. Our findings show that, in the coagulating gland of the male rat, the two secretory proteins studied are processed in parallel, but at completely different intracellular pathways. They are released via different extrusion mechanisms. Transglutaminase is synthesized outside the endoplasmic reticulum, reaches the apical cell pole by free flow in the cytoplasm, and is released via apocrine blebs, the membranes of which appear to be derived from the apical plasma membrane. The protein 115 K, on the other hand, follows the classic route, being synthesized within the cisternae of rough endoplasmic reticulum, subsequently glycosylated in the Golgi apparatus, and released in a merocrine fashion. The mutual exclusion of the two secretory pathways and the regulation of the alternative release mechanism are still unresolved issues.     

Seasonal changes in the fine structure of the accessory sex gland in the mole (Talpa europaea)

The mole has a single pair of accessory sex organs with features of both the prostate and the seminal vesicle, for which the term prostate gland is not appropriate. Seasonal changes occuring in this gland were related to four periods: a) the quiescence period, b) the maturation period, c) the active period and d) the involution period. During the quiescence period the cuboidal epithelial cells display a quasi-embryonic fine structure and are sparse in cytoplasmic organelles, but rich in glycogen and lipopigment. With the onset of sexual activity glycogen and lipopigment disappear and the rough endoplasmic reticulum as well as the Golgi apparatus begin to proliferate. The fully active gland is lined by a low epithelium with parallel stacks of rough endoplasmic reticulum, a large Golgi apparatus and several lysosomes and secretory granules. In the involution period the gland collapses and the epithelial cells are eliminated by hetero- and autophagic processes. During this period a great number of presumably endocrine cells were observed. The results were compared with findings in experimental studies and those on postnatal development of accessory sex glands in laboratory animals.     

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.     

Secretory cell activity in the hamster seminal vesicle following castration. A morphometric ultrastructural study

The ultrastructure of hamster seminal vesicle epithelium was studied 7, 14, 21 and 28 days after castration using a stereological approach. The results show that castration promotes epithelial reorganization, mainly characterized by reduced epithelial cell size and number, decreased rough endoplasmic reticulum and Golgi complex, increased lysosomes and lipid droplets, increased apical secretory granule size and number, and increased intracellular secretory products per average epithelial cell. It is concluded that after testosterone withdrawal the secretory activity of hamster seminal vesicle epithelial cells, although reduced, is not abolished, and that exocytosis is relatively more reduced than secretory protein production. We suggest that an extracellular androgen source is responsible for secretory activity not being lost in the epithelial cells of castrated hamster seminal vesicle.     

Cytodifferentiation in the accessory glands of Tenebrio molitor I. Ultrastructure of the tubular gland in the post-ecdysial adult male

The tubular accessory reproductive glands of the male mealworm beetle consist of a secretory epithelium surrounded by a thin muscular sheath. Each columnar secretory cell is divisible into three zones: basal which is adjacent to the muscle layer and contains rough endoplasmic reticulum and Golgi, intermediate, which contains endoplasmic reticulum and Golgi zones in the immature gland and is filled with secretory vesicles in the mature gland, and apical. Maturation also involves proliferation and organization of the rough endoplasmic reticulum in the basal and intermediate zone. The process appears to be complete at four days after ecdysis. Parallels with other insect glands and with the mammalian prostate are striking.     

Intracellular localization of Prostatic Binding Protein (PBP) in rat prostate by light and electron microscopic immunocytochemistry

Extra- and intracellular distribution of Prostatic Binding Protein (PBP) was studied in the different genital organs of the male rat by immunocytochemistry at the light and electron microscopic levels. PBP was extracted from cytosols of rat ventral prostate and used for immunization of rabbits. The specificity of the antiserum raised was tested by "western blotting" and immunoelectrophoresis. From the different fixatives tested for optimal structural and antigenic preservation of the ventral prostate a mixture containing 2.5% paraformaldehyde, 0.5% glutaraldehyde and 0.5% CaCl2 in cacodylate buffer, 0.05 M, pH 7.3 was selected. Using the immunofluorescence technique and the unlabeled antibody enzyme method PBP-immunoreactivity was detected at the light microscopic level in the luminal secretions of the ventral prostate. No reaction was observed with the seminal vesicle, the coagulating gland, the dorsal and lateral prostates, the epididymis and the testis. Intracellular secretory granules reacting with PBP antiserum were exclusively found in the secretory cells of the ventral prostate. Insufficiently fixed cells showed a diffuse generalized reaction of the cytoplasm indicating a leakage of the antigen from the secretory granules. Such artifacts were common in tissue sections processed with the preembedding-staining procedure. At the ultrastructural level therefore mostly the postembedding staining method was performed using both the unlabeled antibody enzyme method and the ferritin-labeled immunoglobulin technique in osmicated, Epon-embedded tissue. Labeling with either method was intense in the secretory granules and the condensing vacuoles, while the labeling density of the rough endoplasmic reticulum and the Golgi cisternae was in the background range. Castration experiments showed that secretory material displaying PBP immunoreactivity was retained within the acinar lumen of the gland for several days after castration, but was absent from most secretory cells already by four days after castration. Immunocytochemistry of PBP therefore is a very sensitive method for analysing the secretory activity and its androgen dependence of the prostate of the rat.     

小地老虎雄蛾中胚层生殖道和附腺的细胞结构和分泌功能

通过光镜、电镜及组织化学等方法,研究了小地老虎生殖前期雄蛾中胚层生殖道和附腺的腺细胞结构和分泌功能,以及与精子形态和数量变化的关系,结果表明：(1)以缢缩位置、解剖形态、细胞结构、分泌方式、精子形态变化和数量变动为依据,将中胚层生殖道划分为修精囊、输精管、贮精囊、精包腺1～5段等8个区段;(2)中胚层生殖道和附腺具有相同的组织层次,自内向外分为单细胞上皮层、底膜、肌肉层和围膜等4层,但缺少表皮质内膜;(3)中胚层生殖道和附腺的腺细胞具有旺盛的合成和分泌蛋白质的能力,主要有内质网型和液泡型两种,前者有发达的粗面内质网和高尔基体,后者具有致密的核糖体和分泌泡;至少有4种分泌方式：即颗粒顶泌、液泡顶泌、胞质局泌和胞间分泌;修精囊、贮精囊、雄性附腺、精包腺1段的顶泌物为糖蛋白性质(PAS阳性)、局泌物为非糖蛋白性质(PAS阴性)。     

Ultrastructural and cytochemical studies of the effects of prolactin on the lateral prostate and the seminal vesicle of the castrated guinea pig

Summary Administration of ovine prolactin to castrated guinea pigs for 2 weeks induced hypertrophy of secretory cells in the lateral prostate when compared with the castrated controls. This was accompanied by an apparent increase in the number of profiles of granular endoplasmic reticulum and well developed Golgi complexes with dilated cisternae. An increase in the number of low-contrast electron-dense secretory granules was observed 4 weeks after prolactin treatment. In the seminal vesicle, dilatation and degranulation of granular endoplasmic reticulum and an apparent decrease in the number of secretory granules were observed 4 weeks after prolactin administration. Following castration and 2 weeks after prolactin treatment, thiamine pyrophosphatase (TPPase)-reaction product was mainly confined to 1–2 trans cisternae of the Golgi complexes in secretory cells of the lateral prostate and the seminal vesicle. In both glands, a reduction of TPPase activity was observed 2 weeks following prolactin administration, and the reaction product was totally absent after prolonged treatment for 4 weeks. The present study has provided morphological evidence that prolactin is capable of stimulating the secretory function of the lateral prostate while exerting some inhibitory effects on the seminal vesicle of the castrated guinea pig. In both glands, TPPase activity, and hence the process of glycosylation was inhibited after prolactin administration. The results from radioimmunoassay indicated that the action of prolactin on these glands could be a direct effect and not mediated through testosterone.     

Ultrastructure and function of the seminal vesicle of Bittacidae (Insecta: Mecoptera)

The fine structure of the seminal vesicle and reproductive accessory glands was investigated in Bittacidae of Mecoptera using light and transmission electron microscopy. The male reproductive system of Bittacidae mainly consists of a pair of testes, a pair of vasa deferentia, and an ejaculatory sac. The vas deferens is greatly expanded for its middle and medio-posterior parts to form a well-developed seminal vesicle. The seminal vesicle is composed of layers of developed muscles and a mono-layered epithelium surrounding the small central lumen. The epithelium is rich in rough endoplasmic reticulum and mitochondria, and secretes vesicles and granules into the central lumen by merocrine mechanisms. A pair of elongate mesodermal accessory glands opens into the lateral side of the seminal vesicles. The accessory glands are similar to the seminal vesicle in structure, also consisting of layers of muscle fibres and a mono-layered elongated epithelium, the cells of which contain numerous cisterns of rough endoplasmic reticulum and mitochondria, and a few Golgi complexes. The epithelial cells of accessory glands extrude secretions via apocrine and merocrine processes. The seminal vesicles mainly serve the function of secretion rather than temporarily storing spermatozoa. The sperm instead are temporarily stored in the epididymis, the greatly coiled distal portion of the vas deferens.     

The architecture of the accessory reproductive glands of the male desert locust. 5: ultrastructure during maturation

The accessory reproductive glands of the male desert locust were studied with the electron microscope from the time of adult emergence until full sexual maturity was attained (10 days). Observations on the changes in the nucleus and the cytoplasm of the glandular epithelium, particularly those organelles involved in the elaboration of cell secretions, were made during the maturation of the accessory glands. Cells of gland 16 (the functional seminal vesicle) exhibit numerous ribosomal aggregates, have a concentration of mitochondria along their luminal surface, and show some secretory activity, even at the time of adult emergence. On the other hand, all other accessory glands show very little development of the secretory apparatus (rough-surfaced endoplasmic reticulum, polyribosomes, Golgi elements, elaborate mitochondria, etc.) until after the fifth adult day. These changes parallel changes in the secretory activity of the adult corpus allatum; and it is suggested that the corpus allatum hormone regulates the maturation of the truly glandular accessory glands of the locust, but not that of the functional vesicle.     

Effect of gossypol on pituitary reproductive axis: ultrastructural and biochemical studies

Oral administration of gossypol induced sterility in male rats by 10 weeks, at a dose of 15 mg/kg body weight/day. The pituitary FSH gonadotroph cells showed dilated endoplasmic reticulum and accumulation of secretory granules in the cytoplasm. LH cells were degranulated. The Leydig cells showed enhanced synthetic activity. There was no change in testis weight and testicular RNA, lipids and cholesterol in the treated group while significant increase was observed in DNA content. Testicular sialic acid content decreased significantly over controls. The Sertoli cells, spermatogonia, spermatocytes and early spermatids were not affected after the treatment. The weights of prostate, seminal vesicle were recorded normal and there were no ultrastructural variations. The levels of acid and alkaline phosphatase and RNA in prostatic tissue were insignificant as compared with controls. However, DNA content of prostate gland showed a significant increase. Sialic acid of seminal vesicle + coagulating gland were within the control range. A marked reduction in fructose values from the same organ was noted.     

Exit of nonglycosylated secretory proteins from the rough endoplasmic reticulum is asynchronous in the exocrine pancreas

The path and synchrony of intracellular transport of 12 secretory proteins of the guinea pig exocrine pancreas have been studied in pulse-chase amino acid labeling experiments by quantitative analysis of the individual proteins recovered in subcellular fractions and extracellular samples. Protein fractionation was accomplished by two-dimensional isoelectric focusing/SDS-gel electrophoresis. Use of a double-label protocol allowed correction of the data on a protein-by-protein basis for leakage and adsorption artifacts which accompany tissue homogenization. All the labeled secretory (pro)enzymes, including their isoenzymic forms, were recovered in rough microsomal, Golgi-enriched and granule fractions during their transport to the cell surface. However, major asynchrony was observed at four levels: exit from the rough endoplasmic reticulum; transit through the Golgi complex; entry into granules; and discharge from the cell. Rapid transport rates were observed for trypsinogen, chymotrypsinogen 2, procarboxypeptidase A2, and lipase 2. Slow transport rates were observed for amylase and procarboxypeptidase B. In the presence of carbamylcholine or cholecystokinin stimulation, the times required for 40% discharge of labeled chymotrypsinogen 2, trypsinogen, amylase, and procarboxypeptidase B were 98, 102, 148, and 180 min, respectively. Transport rates did not correlate with isoelectric point, molecular weight, or the presence of carbohydrate. These data suggest that interactions occur within the rough endoplasmic reticulum, either between secretory (nonglyco)-proteins themselves or between such proteins and the cisternal face of the rough endoplasmic reticulum.     

Stereology and ultrastructure of the seminal vesicle of C57/BL/6J mice following chronic alcohol ingestion

Excessive alcohol consumption causes metabolic changes and pathologic alterations in testes and accessory sex organ in different animal species. The aim of the present study was to evaluate the macroscopic, histologic and ultrastructural alterations provoked by chronic ingestion of different ethanol concentrations over increasing periods of time on the secretory epithelium of the seminal vesicle of C57/BL/6J mice in using stereological methods. Sixty male adult mice were divided into three experimental groups: Control, Alcoholic 25% and Alcoholic 35%, respectively, receiving tap water and tap water containing ethanol diluted to 25 and 35 degrees Gay Lussac. All mice were fed with the same solid diet. After 150 and 250 days of treatment the animals were sacrificed and the seminal vesicles were collected and processed for light and transmission electron microscopy. The cellular, cytoplasmic and nuclear volumes and the area density of autophagic and secretory vacuoles were measured. The histologic alterations observed in the alcoholic mice consisted of a reduction in epithelial size and cell volume, with maintenance of the same nuclear and cytoplasmic ratio as verified in the control groups. The ultrastructural alterations were: increased density of dense body area, decreased density of secretory granule area, and dilated rough endoplasmic reticulum and Golgi cisternae. We conclude that chronic ethanol ingestion causes depleting morphologic alterations in the epithelial cells of the seminal vesicle and negatively affects the secretory process of this gland.     

Immunocytochemical localization of seminal proteins in salivary and lacrimal glands of the rat

Antibodies against 10 different secretory proteins from the accessory sex glands of the male rat were used for immunohistochemical studies of salivary and lacrimal glands from intact and castrated rats, at the light- and electron-microscopic levels. In the parotid gland, secretory acinar cells showed immunoreactivity with antibodies against prostatic binding protein, cystatin-related peptide and acid phosphatase (isoenzyme pI 8.0; 5.6) typical of ventral prostate, and seminal vesicle secretion VI. Western blotting analysis indicated that immunoreactivity against prostatic binding protein was attributable to a subunit, presumably C3. Acid phosphatase pI 5.6 showed a molecular weight of 66 kDa, which is at variance with the prostatic form. Immunoreactivity for secretory transglutaminase, derived from the coagulating gland, was restricted to myoepithelial and stromal cells. In castrated animals, the immunoreactivity of acinar cells was reduced to the background level, whereas stromal transglutaminase immunoreactivity was unaltered. The distribution pattern of immunoreactivity for the proteins mentioned was almost identical in the lacrimal gland. Significant differences were however observed in the immunoreactivity of the inframandibular gland, where serous glandular cells were non-immunoreactive for seminal proteins, with the exception of acid phosphatase isoenzyme pI 8.0. Granules present in the convoluted granular ducts were immunoreactive particularly for acid phosphatase (isoenzyme pI 5.6)but much less for cystatin-related peptide; immunoreactivity was reduced after castration. The straight portion of the inframandibular duct system was immunoreactive for transglutaminase, but no influence of castration was visible. The distribution of immunoreactivity for seminal proteins present in the salivary and lacrimal glands and the pronounced androgen-dependence of their expression point to functional relationships of the respective proteins at both glandular sites.     

Anatomy and ultrastructure of the salivary gland in the thorax of the honeybee worker,Apis mellifera (Insecta,Hymenoptera)

Summary The thoracic salivary gland of the worker honeybee was investigated by dissection, light microscopy, scanning electron microscopy, and transmission electron microscopy. The glands are paired and each lateral half consists of two parts, a smaller external and a larger internal lobe. The lobes are composed of densely packed secretory tubes and ducts, the tubes of which often show ramifications. A reservoir is packed within the anterior medial part of the gland. The secretory tubes are composed of two types of cells, secretory cells, which are most frequent, and parietal cells. Secretory cells are characterized by a basal labyrinth, abundant rough endoplasmic reticulum, dark secretory vesicles, light vesicles of different sizes, and apical microvilli. Parietal cells are smaller and have a characteristically lobed nucleus and no secretory vesicles. Between the cells there are intercellular canaliculi. In the center of each tube there is an extracellular space with a central cuticular channel. The abundance of rough endoplasmic reticulum and the rare occurrence of smooth endoplasmic reticulum implies a saliva with proteins but rarely with pheromones. Between the secretory tubes there are frequently neuronal profiles which are partly in contact with the secretory cells. Thus a nervous control of this gland is, in contrast to previous investigations, clearly demonstrated. The axonal endings contain dark neurosecretory vesicles as well as light synaptic vesicles. Large parts of the glands are surrounded by a thin tissue sheath which has a smooth surface towards the secretory tubes and shows irregular protrusions towards the outer side. This sheath is considered to be a tracheal air sac, and due to its large extension is probably of importance for the hemolymph flow in the thorax.     

The Golgi complex in the esophageal mucous glands of the newly hatched chick

The general morphology of the mucous gland cell and the nature of the secretory granule in esophageal glands of the newly hatched chick have been described. Lightly basophilic supporting cells, attached to secretory cells by desmosomes and containing tonofilaments, are located on the basal lamina. Electron microscopic studies showed a morphological polarity of the Golgi complex which suggests that mucous precursors are transported from other sites within the cell to the Golgi complex for further packaging into secretory granules. Finally, acid mucopolysaccharides (AMPS) were specifically stained using the Thorotrast technique and not detected in the rough endoplasmic reticulum, the transitional elements, or in the lamellae at the forming face of the Golgi complex. Conversely, AMPS are found in the vicinity of the mature face of the Golgi complex, and in the secretory granules. The acquisition of cytochemical reactivity for AMPS within the Golgi complex is discussed.     

The ultrastructure of the accessory sex organs of the male rat

Summary The seminal vesicles and the coagulating gland of the rat were studied 2, 3, 5, 7 and 21 days after castration. The major changes within the seminal vesicles were primarily formation of whorls of the rough endoplasmic reticulum (RER), followed by a general atrophy with a numerical reduction of the RER-profiles, and with general simplification of the cytoplasm due to loss of the organelles. It was a gradually reduction of secretion granules, diminution of the Golgi apparatus, formation of pigment bodies and autophagic vacuoles. Lipid droplets were observed in the basal cytoplasm of the epithelial cells. In the coagulating gland, similar changes occurred within the Golgi area and the lysosome complex. On the other hand, cisternae of the basal endoplasmic reticulum tended to persist in many cells. The similarity in response strongly suggests that the pathogenetic mechanisms are similar in both organs, i.e. atrophy due to deprivation of the androgenic stimulus. The deprivation of androgen gave rise to an inflammatory-like process with infiltration of lymphocytes and macrophages. The increased number of macrophages may indicate that they contribute in some way to the involution of the prostate by removing the material in the autophagic vacuoles.     

The arrangement of the Golgi complex beads is not controlled by the cytoskeleton

Exposure of insect fat body to treatments which disrupt microtubules (colchicine, vinblastine sulfate and cold treatment) blocks intracellular transport between the Golgi complex and the plasma membrane but does not affect Golgi complex bead rings or transport from rough endoplasmic reticulum to the Golgi complex. Drugs which disrupt microfilaments (cytochalasins B and D) do not affect the bead rings or intracellular transport of secretory proteins at any level. Thus, intracellular transport between the rough endoplasmic reticulum and the Golgi complex and the arrangement of the beads in rings are both independent of the cytoskeleton. The ring arrangement is presumably maintained by interconnection(s) with rough endoplasmic reticulum membrane.