Ultrastructure of pancreatic exocrine cells of the rat during starvation

Ultrastructural changes of the pancreatic exocrine cells after 3, 7, 14, 21, 28, 35 and 42 days of starvation were observed in male rats aged from 16 to 18 months weighing between 600 and 700 grams. The number of zymogen granules after starvation decreased to less than about 70 per cent of that of the control. Changes in the rough endoplasmic reticulum were hardly seen up to 14 days of starvation as compared with the control, but were observed in the apical and basal cytoplasm of the cell from 21 days after starvation. Particularly in 35- and 42-day starved rats, the rough endoplasmic reticulum was frequently shortened and dilated, and changed to disorganized membranous structures. The lysosomes in the apical cytoplasm of the cell gradually increased in number after starvation, and contact or fusion between the zymogen granules and lysosomes (viz, so-called crinophagy) was often seen at 35 and 42 days of starvation. Large autolysosomes especially those containing zymogen granules and rough endoplasmic reticulum were also marked in the basal cytoplasm of the cell after 35 and 42 days of starvation. Alterations in the basal cytoplasm of the cell appeared later than those in the apical cytoplasm. It was considered that, owing to its role in protein synthesis, the basal cytoplasm of the pancreatic exocrine cells in starved rats might be protected as far as possible during long-term starvation.     

Effects of pinealectomy on the ultrastructure of the golden hamster parathyroid gland

Ultrastructural changes of the parathyroid glands of pinealectomized golden hamsters were investigated. The main changes in the parathyroid glands 1 hour and 1 day after pinealectomy compared with the control and sham-operated groups were an increase of the Golgi complexes, cisternae of the granular endoplasmic reticulum and large vacuolar bodies. In addition, many chief cells contained numerous prosecretory granules in the Golgi areas and many secretory granules in the peripheral cytoplasm. The morphology of the parathyroid glands 7 and 30 days after pinealectomy resembled that of the control parathyroid glands. These results suggest that pinealectomy affects the secretory activity of the parathyroid gland.     

A morphometric study of the secretory process in the endocrine pancreas of the foetal rat

The present study concerns pancreatic beta cells from rat foetus at 18, 19 and 21 days of gestation. On micrographs, the cytoplasm of beta cells was subdivided into 3 zones: one zone corresponding roughly to the cell web, a second zone just underlying the cell web, and a third zone comprising the remaining cytoplasm. The secretory granules present in each zone were counted; in the cell web, granules fused with plasma membrane were counted separately. During later foetal stages the increase in the frequency of granule to plasma membrane fusions parallels the increase in blood insulin levels, and the total number of granules in beta cells increases in parallel with the pancreatic insulin content. Therefore, as the beta cell matures, both secretion and biosynthesis of insulin increase sharply. The observed changes in the distribution of the granules in the different zones of the cytoplasm with the foetal age suggests that the cell web controls the access of the granules to the plasma membrane. The morphometric technique used allows a direct determination, at the cellular level, of even small variations in exocytosis-mediated secretory discharge and suggests a regulatory role of the cell web.     

Effects of isoproterenol on the fine structure of the hamster parathyroid gland

Ultrastructural changes of the parathyroid glands of isoproterenol-treated golden hamsters were investigated. Many chief cells in the parathyroid glands after 5 and 10 minutes of administration of isoproterenol contain well-developed Golgi complexes and granular endoplasmic reticulum, numerous prosecretory granules, and many secretory granules in the peripheral cytoplasm as compared with the control animals. Many chief cells in the parathyroid glands after 1, 3, 6 and 12 hours of administration have poorly-developed Golgi complexes, granular endoplasmic reticulum, many secretory granules and numerous lipid droplets as compared with the control animals. The morphology of the parathyroid gland after 30 minutes and 24 hours of administration resembles that of the control animals. It is considered that isoproterenol affects the secretory activity of the parathyroid gland.     

Ultrastructure of endocrine pancreatic granules during pancreatic differentiation in the grass snake,Natrix natrix L. (Lepidosauria,Serpentes)

We used transmission electron microscopy to study the pancreatic main endocrine cell types in the embryos of the grass snake Natrix natrix L. with focus on the morphology of their secretory granules. The embryonic endocrine part of the pancreas in the grass snake contains four main types of cells (A, B, D, and PP), which is similar to other vertebrates. The B granules contained a moderately electron‐dense crystalline‐like core that was polygonal in shape and an electron‐dense outer zone. The A granules had a spherical electron‐dense eccentrically located core and a moderately electron‐dense outer zone. The D granules were filled with a moderately electron‐dense non‐homogeneous content. The PP granules had a spherical electron‐dense core with an electron translucent outer zone. Within the main types of granules (A, B, D, PP), different morphological subtypes were recognized that indicated their maturity, which may be related to the different content of these granules during the process of maturation. The sequence of pancreatic endocrine cell differentiation in grass snake embryos differs from that in many vertebrates. In the grass snake embryos, the B and D cells differentiated earlier than A and PP cells. The different sequence of endocrine cell differentiation in snakes and other vertebrates has been related to phylogenetic position and nutrition during early developmental stages.     

Ultrastructure of the preparative phase of cell death in the larval fat body of Drosophila melanogaster

Progressive changes in the ultrastructure of the larval fat body of Drosophila melanogaster were studied during the third instar. In addition to electron microscopy, light microscopy and morphometric stereology were employed to evaluate the tissue at five 12-hr intervals: 48, 60, 72, 84, and 96 hr after hatching from the egg. Lipid and glycogen were found stored throughout the instar, whereas protein is stored in the form of cytoplasmic granules mainly during the final 24 hr. The cells increased in cross-sectional area, and there was a concomitant increase in the relative amounts of these substances. Based on morphological characteristics there were three types of protein granules which we called dense granules (D), heterogeneous granules (H), and autophagic vacuoles. The morphology, size range, time of appearance, and changes in frequency of these granules suggested that the H type arose from D granules, and that the autophagic vacuoles were derived from D and H types. Morphological evidence indicated D granules have the unusual characteristic of forming in the intercellular space before entering the cytoplasm.     

Occurrence of unusual heterogeneous lipid-containing granule-storing cells in the main excretory duct epithelium of the male mouse submandibular gland

Summary The fine structure of the main excretory duct epithelium of the male mouse submandibular glands was investigated by scanning and transmission electron microscopy. Three principal cell-types were observed: type I and II, and basal cells. This epithelium was characterized by the presence of intercellular canaliculi. Type-I cells were the most numerous. They had an abundance of mitochondria, well-developed Golgi apparatus, a few electron-lucent lipid-containing granules and poorly developed basal infoldings. These cells were also characterized by many glycogen granules throughout the cytoplasm and abundant smooth endoplasmic reticulum in the apical cytoplasm. Type-II cells were the second most numerous. Their most characteristic feature was the presence of abundant heterogeneous lipid-containing granules having acid phosphatase activity at the periphery. They were concentrated in the infra- and supranuclear cytoplasm. The granules may be derived from mitochondrial transformation and seem to be a special kind of secondary autolysosome. Type-II cells also contained abundant mitochondria throughout the cytoplasm, much smooth endoplasmic reticulum in the apical cytoplasm, a well developed Golgi apparatus adjacent to the heterogeneous lipid-containing granules and no basal infoldings. Basal cells were situated adjacent to the basal lamina. They had a large nucleus and the cytoplasm was filled with glycogen granules.     

抗体形成细胞发育的免疫电镜研究

The pre-embedding immunoelectron microscopic method was used to study the development of antibody-producing cells in the guinea pig popliteal lymph nodes of 2, 3, 5, 8 and 10 days after a second challenge with horseradish peroxidase. The results indicated that the antibody activity was located in the perinuclear space, the endoplasmic reticulum and Golgi complex. According to the cellular developmental stages and the characteristics of distribution of the antibody activity, the antibody-producing cells (APC) were divided into four types: (1) Type I cells (lymphocytes) exhibited many positive granules throughout the cytoplasm; (2) Type II cells (proplasmacytes) contained many positive granules and positive short bars, some of them were parallel; (3) Type III cells (proplasmacytes) contained numerous parallel positive lamellae in cytoplasm; (4) the parallel lamellae in cytoplasm of type IV cells (plasmacytes) were arranged into a network-endoplasmic reticulum. According to the kinetic change from granules, short bars to parallel lamellae and the network, the results indicated the developmental course of AFC from lymphocytes, proplasmacytes to plasmacytes.     

Effects of melatonin on the ultrastructure of the golden hamster parathyroid gland.

Ultrastructural changes of the parathyroid glands of melatonin-treated golden hamsters were studied. Many chief cells in the parathyroid glands after 1 hour of administration of melatonin contained poorly-developed Golgi complexes associated with a few prosecretory granules and numerous lipid droplets as compared with those of the control animals. The morphology of the parathyroid glands after 5 hours of administration resembled that of the control animals. Many chief cells in the parathyroid glands after 24 hours of administration had well-developed Golgi complexes and cisternae of the granular endoplasmic reticulum, numerous prosecretory granules, a few lipid droplets and many secretory granules in the peripheral cytoplasm as compared with those of the control animals. The ultrastructure of the parathyroid glands after 48 hours of administration was almost similar to that of the control animals. It is considered that melatonin affects the secretory activity of the parathyroid gland.     

Immunocytochemical studies on the pancreatic endocrine cells in the Japanese newt (Cynopus pyrrhogaster)]

Pancreatic endocrine cells were examined by light and electron microscopic immunocytochemistry to discuss the co-localization of peptides in one cell type. A cells were irregular in shape with an occasional long cytoplasmic process, and contained glucagon-immunoreactive granules with various contours. These granules were 160-300nm in diameter with various density, and also immunoreactive to anti-human pancreatic polypeptide (PP) serum. A part of them were further immunoreactive to anti-somatostatin serum. B cells were round to elliptical in shape, and often aggregated around the capillaries. Granules of B cells were round to irregular in shape, 270-410 nm in diameter, and immunoreactive to anti-insulin serum. D cells were irregular in shape with meager cytoplasm, and contained somatostatin-immunoreactive granules. These granules were ovoid or teardrop in shape, 140-250nm in longitudinal diameter, and immunoreactive to both anti-somatostatin and anti-human PP sera. PP cells were round to spindle-shaped, and contained human PP-immunoreactive round granules 150-35nm in diameter. These findings reveal the existence of at least 4 types of endocrine cells secreting glucagon, insulin, somatostatin, and PP, respectively, in the newt pancreas, and suggest the co-localization of some of these peptides in one cell type.     

Fine structure and immunocytochemistry of cells within the endocrine pancreas of larval and adult sea lampreys, Petromyzon marinus L

Immunocytochemistry with protein A-gold and routine electron microscopy were used to identify cell types within the endocrine pancreas of larvae, juvenile adults, and upstream-migrant adults of the sea lamprey, Petromyzon marinus. The larval pancreatic islets are composed only of insulin-immunoreactive B-cells, which are uniform in their fine structure. The cranial and caudal pancreatic tissue in both adult periods contains three cell types: B-cells, somatostatin-immunoreactive D-cells, and a third cell type of unknown content. No glucagon-immunoreactive cells are present in lampreys, but B- and D-cells exist in equal numbers in the pancreatic tissue of adults. The B-cells of adults have a fine structure similar to those in larvae. D-cells have secretory granules that are distinctly different from those both in B-cells and in the third cell type. Although B- and D-cells in lamprey pancreatic tissues have a basic morphological similarity to these cells in other vertebrates, their granules are generally of smaller dimensions. The inclusion of granules within large pleomorphic bodies in many D-cells indicates that granule turnover is common. Immunocytochemistry will be a useful tool for showing the relationship between the cells in the degenerating bile ducts and those of the developing adult pancreas.     

Identification of pancreatic glucagon cells in the cartilaginous fish Scyliorhinus canicula by ultrastructural immunocytochemistry

The ultrastructural localization of glucagon in the presence of Scyliorhinus canicula was investigated. We used a post-embedding immunoelectron microscopy method on pancreatic samples fixed in glutaraldehyde and osmicated before embedding. Contrasting with uranyl acetate and lead citrate was also performed after immunolabelling, but best results were obtained with uranyl acetate only. Glucagon-like immunoreactivity was located in round granules (300-600 nm) surrounded by a limiting membrane. The matrix varied in electron density and exhibited a dense core surrounded by a less dense mantle. The granules were seen in two different cell types, which differed in the electron density of their cytoplasm. Glucagon-immunoreactive cells were the largest pancreatic cells types and were often localized near somatostatin-containing cells.     

Ultra-structural study of insulin granules in pancreatic β-cells of db/db mouse by scanning transmission electron microscopy tomography

Insulin granule trafficking is a key step in the secretion of glucose-stimulated insulin from pancreatic β-cells. The main feature of type 2 diabetes (T2D) is the failure of pancreatic β-cells to secrete sufficient amounts of insulin to maintain normal blood glucose levels. In this work, we developed and applied tomography based on scanning transmission electron microscopy (STEM) to image intact insulin granules in the β-cells of mouse pancreatic islets. Using three-dimensional (3D) reconstruction, we found decreases in both the number and the grey level of insulin granules in db/db mouse pancreatic β-cells. Moreover, insulin granules were closer to the plasma membrane in diabetic β-cells than in control cells. Thus, 3D ultra-structural tomography may provide new insights into the pathology of insulin secretion in T2D.     

Chloride secretion in the submandibular gland of adult and early postnatal rats studied by X-ray microanalysis

Submandibular acinar cells of 1-day-old, 7-day-old, and adult rats were analyzed with X-ray microanalysis after stimulation with carbachol for different time periods (2–7 min). In unstimulated animals, marked differences in elemental content between compartments could be observed: secretory granules had a higher Ca and lower P and K content than other cell compartments. Comparison between different age groups showed significant differences for Ca, which increased with age in all compartments; Mg increased with age in the secretory granules and the apical cytoplasm. Only the glands from adult animals showed a significant effect of cholinergic stimulation: a transient decrease in Cl and K. The Cl concentration in the secretory granules decreased to 60% of the control value, which suggests that the granules release Cl upon stimulation. In young animals, no or little change in elemental distribution was observed after stimulation. This may indicate that Cl-secretion mechanisms are much less prominent in young animals. The ultrastructure of submandibular secretory granules depends on the preparation method: condensed and electrondense in freeze-substituted unfixed tissue, decondensed and more translucent in aldehyde-fixed tissue. This may indicate that the granules can transport water, and swell during the process of aldehyde fixation.     

Response of exocrine pancreatic secretion to CCK in adrenalectomized rats

A study was made of the effect of adrenalectomy over different periods of time (6, 15 and 21 days) on exocrine pancreatic secretion in the rat in basal conditions and under stimulation with CCK. It was observed that adrenalectomy does not alter the rate of pancreatic flow but the response capacity to CCK is depressed. The secretion of total protein and amylase decreases significantly after sixth day, reaching the lowest levels after 21 days. Despite this, after 6 days the adrenalectomized rats showed the same capacity of response to CCK as the non-adrenalectomized animals, while after longer periods of time (15 and 21 days) the response to CCK was inhibited. The fact that the lack of glucocorticoids prevents the maturation of zymogen granules seems to be the main reason why the acinar cells do not increase protein secretion in response to CCK at 15 and 21 days after adrenalectomy. It is concluded that the sensitivity of exocrine pancreas to CCK and the amount of zymogen granules in the acinar cells decrease as a function of the time over which the animals are deprived of glucocorticoids.     

Effects of VIP on the regulation of mucin secretion in cultured human pancreatic cancer cells (Capan-1)

Summary The effects of Vasoactive intestinal peptide (VIP) on mucin secretion in the pancreatic cancer Capan-1 cell line were studied by Enzyme-linked-immunosorbent-assay (ELISA), and by light and electron microscopy using immunocytological methods. During the exponential growth phase, mucins were accumulated in the cytoplasm of cells and slowly exocytosed. In contrast, there was enhanced exocytosis of mucins during the stationary phase when the cells were well-polarized. Moreover, during this phase, VIP induced a dose-dependent rise in mucin content in the extracellular medium. The reaction with anti-MI monoclonal antibodies, which recognize specifically the peptide core of gastric mucins, showed an accumulation of secretion granules near the apex of well-polarized cells together with fusion of the granule and plasma membranes after VIP stimulation. Moreover, mucin exocytosis was stimulated by Pituitary adenylate cyclase activating polypeptide (PACAP) and secretin. It was also increased after forskolin treatment suggesting that this mechanism was cAMP-dependent. Our results suggested that exocytosis of mucins could be under the control of VIP in pancreatic duct cells of the Capan-1 cell line.     

Quantitation of ultrastructural changes in mouse pancreatic acinar cells caused by foreign serum

Quantitative changes in the pancreatic acinar cell organelles were studied in BALB/c mice injected with 1.0 ml fresh rabbit serum intraperitoneally. Groups of 5 mice were killed at 0, 1, 3, 6 and 12 h after the serum injection. Pancreatic tissue was processed for electron microscopy by glutaraldehyde and osmium tetroxide fixation and Epon embedding. The proportions of acinar cell cytoplasm (volume fractions) occupied by zymogen granules, granular endoplasmic reticulum, Golgi apparatus, mitochondria and lysosomes (including autophagosomes) were determined by the point counting method from electron micrographs. The volume fraction of lysosomes increased during the first 3 h and remained markedly elevated up to 12 h. The volume fractions of zymogen granules increased from 12 to 28% in 12 h. It was concluded that the secretory mechanism of pancreatic acinar cells was injured by the foreign serum. The injury caused accumulation of zymogen granules and increased autophagic activity in the acinar cells.     

Twenty four hour variations in subcellular structures of rat pancreatic islet B-, A- and D-cells,and of portal plasma glucose and insulin levels

Summary Variations in stomach weights, plasma glucose and insulin levels in the portal vein, and in subcellular structures of rat pancreatic islet B-, A- and D-cells were examined over 24 h. The variation in stomach weights paralleled plasma glucose levels, indicating that the levels may be influenced by intestinal glucose absorption. Plasma insulin levels increased from the late dark to the early light periods, whereas they decreased from the late light to the early dark periods. The variation in plasma insulin levels was in the opposite sense to that in the relative numbers of B-cell granules. The decrease in the relative numbers of A-cell granules occurred between the late dark and early light periods. The relative numbers of D-cell granules decreased before and after the decreases in B- and A-cell granules. The variation in D-cell granules appears to correspond to the inhibitory effect of somatostatin. The relative amounts of rough endoplasmic reticulum (rER) in each cell varied in a reciprocal manner to those of B-cell granules. Moreover, the variation in plasma insulin levels coincided with variations in rER of hepatocytes and pancreatic acinar cells. The changes in rER of each cell may correlate with the trophic effect of insulin.     

Morphometric studies of pancreas islets of rats after subphrenic vagotomy]

Analysis of the morphometric data showed that 7 days after vagotomy significant changes ocurred in the pancreatic islets, mainly as a result of variation in B cells. At later periods (45 and 90 days after surgery) normalization of some morphofunctional parameters is noted, however, absolute restoration is not achieved.     

Kinematographische Dokumentation einer amitotischen Zellteilung

Summary Identification of argyrophilic cells, in pancreatic islets of normal rabbits, is accomplished by light and electron microscopy in osmium-fixed plastic-embedded tissues.Fixative, pretreatment and pH of silver nitrate solution were essential for the light microscopic study to reveal argyrophilic cells in osmium-fixed plastic-embedded pancreatic islet tissue. The best result was obtained with Dalton's osmium fixation and buffered silver nitrate methanamine solution at pH 9.O. The cytoplasmic granules of argyrophilic cells generally are densely packed but some of the cells show only sparse silver impregnated granules in the cytoplasm. Occasionally there are some non-argyrophilic granular cells in which, after silver impregnation, the cytoplasm appears clear. There are three kinds of cells in the pancreatic islets, i.e., argyrophilic granular cells, non-argyrophilic granular (clear) cells, and beta cells (situated centrally in the islet and stained light yellow in silver impregnated sections).The cells known as argyrophilic cells in light microscopy can be identified as alpha cells in electron micrographs by comparison of consecutive sections of the same cell.The author would like to express his appreciation to professor Roy C. Swan for his generous guidance.