scholarly journals Effect of taxol on secretory cells: functional, morphological, and electrophysiological correlates.

1985 ◽  
Vol 100 (6) ◽  
pp. 1863-1874 ◽  
Author(s):  
J Thuret-Carnahan ◽  
J L Bossu ◽  
A Feltz ◽  
K Langley ◽  
D Aunis
Keyword(s):  
Chromaffin Cells ◽  
Secretory Activity ◽  
Ultrastructural Level ◽  
Secretory Cells ◽  
Patch Clamp Technique ◽  
Cell Axis ◽  
Ultrastructural Studies ◽  
Microtubule Polymerization ◽  
Long Term Effect

The effect of 0.5-1.0 microM taxol, a potent promoter of microtubule polymerization in vitro, was studied on the secretory activity of chromaffin cells of the adrenal medulla. Taxol was found to have a dual effect: the long-term effect (after a 1-h incubation) of taxol was to induce almost complete inhibition of catecholamine release, whereas after a short incubation (10 min) a massive, nicotine-independent release of catecholamine was produced. From results obtained using the patch-clamp technique to study the Ca++-dependent K+ channels (Ic channels), it was possible to conclude that taxol probably provokes an augmentation of free [Ca++]i in the cytoplasm, values increasing from 10(-8) M at rest to several 10(-7) M. The increased spontaneous release of stored neurohormones and the increased frequency of opening of Ic channels occur simultaneously and could both originate from a rise of [Ca++]i upon taxol addition. Immunofluorescence and ultrastructural studies showed that 13-h taxol treatment of chromaffin cells led to a different distribution of secretory organelles, and also to microtubule reorganization. In treated cells, microtubules were found to form bundles beneath the cell membrane and, at the ultrastructural level, to be packed along the cell axis. It is concluded that in addition to its action on microtubules, the antitumor drug taxol has side effects on the cell secretory activity, one of them being to modify free [Ca++]i.

ULTRASTRUCTURE AND IMMUNOCYTOCHEMISTRY OF THE PARS DISTALIS OF THE FEMALE RAT WHEN GRAFTED UNDER THE KIDNEY CAPSULE AND A PARALLEL STUDY OF THE PLASMA LEVELS OF PROLACTIN

1981 ◽  
Vol 89 (1) ◽  
pp. 157-NP ◽  
Author(s):  
L. I. AGUADO ◽  
G. S. ALVIAL ◽  
E. M. RODRÍGUEZ
Keyword(s):  
Light And Electron Microscopy ◽  
Secretory Activity ◽  
Ultrastructural Level ◽  
Lymphoid Cells ◽  
Female Rats ◽  
Secretory Cells ◽  
Female Rat ◽  
Plasma Prolactin ◽  
Kidney Capsule ◽  
Hormone Content

Two hypophysial partes distales were grafted under the kidney capsule of intact female rats. The plasma prolactin levels 15, 45 and 90 days after the operation were determined. At the same postoperative intervals the grafted glands of some of the operated rats were processed for conventional light and electron microscopy and for the demonstration of prolactin, FSH and LH according to the unlabelled immunoperoxidase procedure. The ultrastructural characteristics of the transplanted secretory cells and the amount and distribution of the immunoreactive material within their cytoplasm were used to evaluate approximately the secretory activity of these cells. Although levels of prolactin in the three experimental groups were significantly higher than those in control rats, a decrease in prolactin level was detected in 71% of the samples taken 45 days after operation. At day 15 the graft was completely surrounded by lymphoid cells whereas at day 45 these cells had invaded the whole graft. In the group sampled at day 90 the graft was free of lymphoid cells. When traced immunocytochemically the three types of cells followed different patterns of evolution after transplantation. Most prolactotrophs were hypertrophied in all groups but, in addition, they underwent a process leading to hyperplasia some time between days 45 and 90 after operation. Syncytial formations which probably correspond to multinucleated prolactotrophs were present only in the group sampled at day 90. The number of LH and FSH cells had decreased in the group at day 45 and by day 90 the former remained scarce but immunoreactive FSH cells were no longer found. At the ultrastructural level clear signs of involution of gonadotrophs and degradation by macrophages were seen in the graft 45 days after operation. The relation between the morphology and hormone content of the graft and hormone content of the plasma is discussed, together with several questions raised by the results. Pituitary transplantation can be used as an experimental model only if the time-dependent changes described here are taken into account.

scholarly journals Multiple Forms of Endocytosis In Bovine Adrenal Chromaffin Cells

1997 ◽  
Vol 139 (4) ◽  
pp. 885-894 ◽  
Author(s):  
Corey Smith ◽  
Erwin Neher
Keyword(s):  
Chromaffin Cells ◽  
Initial Rate ◽  
Secretory Activity ◽  
Patch Clamp Technique ◽  
Intact Cells ◽  
Whole Cell ◽  
Perforated Patch ◽  
Cell Configuration ◽  
Adrenal Chromaffin ◽  
Cell Capacitance

We studied endocytosis in chromaffin cells with both perforated patch and whole cell configurations of the patch clamp technique using cell capacitance measurements in combination with amperometric catecholamine detection. We found that chromaffin cells exhibit two relatively rapid, kinetically distinct forms of stimulus-coupled endocytosis. A more prevalent “compensatory” retrieval occurs reproducibly after stimulation, recovering an approximately equivalent amount of membrane as added through the immediately preceding exocytosis. Membrane is retrieved through compensatory endocytosis at an initial rate of ∼6 fF/s. Compensatory endocytotic activity vanishes within a few minutes in the whole cell configuration. A second form of triggered membrane retrieval, termed “excess” retrieval, occurs only above a certain stimulus threshold and proceeds at a faster initial rate of ∼248 fF/s. It typically undershoots the capacitance value preceding the stimulus, and its magnitude has no clear relationship to the amount of membrane added through the immediately preceding exocytotic event. Excess endocytotic activity persists in the whole cell configuration. Thus, two kinetically distinct forms of endocytosis coexist in intact cells during perforated patch recording. Both are fast enough to retrieve membrane after exocytosis within a few seconds. We argue that the slower one, termed compensatory endocytosis, exhibits properties that make it the most likely mechanism for membrane recycling during normal secretory activity.

scholarly journals The participation of annexin II (calpactin I) in calcium-evoked exocytosis requires protein kinase C.

1991 ◽  
Vol 114 (6) ◽  
pp. 1135-1147 ◽  
Author(s):  
T Sarafian ◽  
L A Pradel ◽  
J P Henry ◽  
D Aunis ◽  
M F Bader
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Chromaffin Cells ◽  
Secretory Activity ◽  
Cytosolic Proteins ◽  
Calcium Dependent ◽  
Kinase C ◽  
Adrenal Chromaffin ◽  
In Cells

Permeabilized adrenal chromaffin cells secrete catecholamines by exocytosis in response to micromolar calcium concentrations. Recently, we have demonstrated that chromaffin cells permeabilized with digitonin progressively lose their capacity to secrete due to the release of certain cytosolic proteins essential for exocytosis (Sarafian T., D. Aunis, and M. F. Bader. 1987. J. Biol. Chem. 34:16671-16676). Here we show that one of the released proteins is calpactin I, a calcium-dependent phospholipid-binding protein known to promote in vitro aggregation of chromaffin granules at physiological micromolar calcium levels. The addition of calpactin I into digitonin- or streptolysin-O-permeabilized chromaffin cells with reduced secretory capacity as a result of the leakage of cytosolic proteins partially restores the calcium-dependent secretory activity. This effect is specific of calpactin I since other annexins (p32, p37, p67) do not stimulate secretion at similar or higher concentrations. Calpactin I requires the presence of Mg-ATP, suggesting that a phosphorylating step may regulate the activity of calpactin. Calpactin is unable to restore the secretory activity in cells which have completely lost their cytosolic protein kinase C or in cells having their protein kinase C inhibited by sphingosine or downregulated by long-term incubation with TPA. In contrast, calpactin I prephosphorylated in vitro by purified protein kinase C is able to reconstitute secretion in cells depleted of their protein kinase C activity. This stimulatory effect is also observed with thiophosphorylated calpactin I which is resistant to cellular phosphatases or with phosphorylated calpactin I introduced into cells in the presence of microcystin, a phosphatase inhibitor. These results suggest that calpactin I is involved in the exocytotic machinery by a mechanism which requires phosphorylation by protein kinase C.

Ultrastructural changes in the rough endoplasmic reticulum and its contents following Brefeldin A treatment of human chondrocytes in vitro

1991 ◽  
Vol 49 ◽  
pp. 256-257
Author(s):  
H. E. Gruber
Keyword(s):  
Endoplasmic Reticulum ◽  
Rough Endoplasmic Reticulum ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Brefeldin A ◽  
Ultrastructural Level ◽  
Human Chondrocytes ◽  
Ultrastructural Changes ◽  
Ultrastructural Studies

The rough endoplasmic reticulum (rER) is now recognized as a major organelle responsible for ensuring that only structurally correct and properly folded proteins are allowed to enter the cellular secretory pathway. We are especially interested in the behavior of the chondrocyte rER since ultrastructural studies of many skeletal dysplasias have revealed that electron dense material accumulates or is not degraded within the rER of chondrocytes from patients. Remodelling of the rER in chick chondrocytes has also been evaluated at the ultrastructural level and the rER found to play a role in procollagen export from the cell. We have utilized normal human chondrocytes grown in culture to investigate the role of brefeldin A, an antiviral antibiotic, which has been shown to primarily block protein transport from the ER to the Golgi complex.

Respiratory tract epithelium in primary culture: concurrent growth and differentiation during establishment

1978 ◽  
Vol 32 (1) ◽  
pp. 269-291
Author(s):  
C.A. Heckman ◽  
A.C. Marchok ◽  
P. Nettesheim
Keyword(s):  
Respiratory Tract ◽  
Secretory Cells ◽  
Basal Cells ◽  
Upper Respiratory Tract ◽  
Squamous Cells ◽  
Ultrastructural Studies ◽  
Term Maintenance

Studies of cellular function in the respiratory tract lining have traditionally been limited by the small tissue mass and functional diversity of the epithelium. Recent improvements in culture conditions have permitted long-term maintenance of epithelial cells derived from the upper respiratory tract of rats. The present study determined the extent to which proliferation and differentiation took place in such epithelial cell cultures. The labelling index obtained after 3H-dThd administration was approximately 100-fold higher than that of the quiescent epithelium in vivo; therefore, a large proportion of the cells were probably in the cycling population. Ultrastructural studies showed that features unique to the specialized mucous secretory cells and ciliated cells were lost rapidly with entry of these cells into the in vitro environment. With long-term maintenance, the cultures were reorganized into a stratified epithelium containing large, squamous, apical cells and small basal cells. The ultrastructural appearance of basal cells in vitro was nearly identical to that of basal cells in vivo. Squamous cells were frequently joined by tight junctions. Because hemicysts originated by detachment of squamous cells from the basal layers but not from adjacent squamous cells, they were considered to indicate stratification in the cultures. The proliferative and differentiative status of the mucociliary epithelium was altered by in vitro conditions, and came to resemble that of early squamous metaplasia in the respiratory tract epithelium.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Probing the ultrastructure of the mitotic and meiotic spindle in eggs: An expeditious approach based on semi-thin (0.25 μm) serial sections

1983 ◽  
Vol 41 ◽  
pp. 552-555
Author(s):  
Conly L. Rieder ◽  
S. Bowser ◽  
R. Nowogrodzki ◽  
K. Ross ◽  
G. Sluder
Keyword(s):  
Small Sample Size ◽  
Small Sample ◽  
Meiotic Spindle ◽  
Serial Sections ◽  
Mitotic Apparatus ◽  
Thin Sections ◽  
Cell Cycles ◽  
Ultrastructural Studies

Eggs have long been a favorite material for studying the mechanism of karyokinesis in-vivo and in-vitro. They can be obtained in great numbers and, when fertilized, divide synchronously over many cell cycles. However, they are not considered to be a practical system for ultrastructural studies on the mitotic apparatus (MA) for several reasons, the most obvious of which is that sectioning them is a formidable task: over 1000 ultra-thin sections need to be cut from a single 80-100 μm diameter egg and of these sections only a small percentage will contain the area or structure of interest. Thus it is difficult and time consuming to obtain reliable ultrastructural data concerning the MA of eggs; and when it is obtained it is necessarily based on a small sample size.We have recently developed a procedure which will facilitate many studies concerned with the ultrastructure of the MA in eggs. It is based on the availability of biological HVEM's and on the observation that 0.25 μm thick serial sections can be screened at high resolution for content (after mounting on slot grids and staining with uranyl and lead) by phase contrast light microscopy (LM; Figs 1-2).

Morphological Observations on the Granule Types in Rabbit Extra-Adrenal Chromaffin Cells.

1975 ◽  
Vol 33 ◽  
pp. 318-319
Author(s):  
Joe A. Mascorro ◽  
Robert D. Yates
Keyword(s):  
Chromaffin Cells ◽  
Sodium Phosphate ◽  
Ganglion Cells ◽  
Ultrastructural Level ◽  
Osmic Acid ◽  
Adrenal Chromaffin Cells ◽  
Potassium Iodate ◽  
Perfusion Fluid ◽  
New Zealand White Rabbits ◽  
Adrenal Chromaffin

Extra-adrenal chromaffin organs (abdominal paraganglia) constitute rich sources of catecholamines. It is believed that these bodies contain norepinephrine exclusively. However, the present workers recently observed epinephrine type granules in para- ganglion cells. This report investigates catecholamine containing granules in rabbit paraganglia at the ultrastructural level.New Zealand white rabbits (150-170 grams) were anesthetized with 50 mg/kg Nembutal (IP) and perfused with 3% glutaraldehyde buffered with 0.2M sodium phosphate, pH 7.3. The retroperitoneal tissue blocks were removed and placed in perfusion fluid for 4 hours. The abdominal paraganglia were dissected from the blocks, diced, washed in phosphate buffer and fixed in 1% osmic acid buffered with phosphate. In other animals, the glutaraldehyde perfused tissue blocks were immersed for 1 hour in 3% glutaraldehyde/2.5% potassium iodate buffered as before. The paraganglia were then diced, separated into two vials and washed in the buffer. A portion of this tissue received osmic acid fixation.

Ultrastructural studies of the relationship between actin filaments and secretory granules

1990 ◽  
Vol 48 (3) ◽  
pp. 458-459
Author(s):  
Ellen Holm Nielsen
Keyword(s):  
Electron Microscopy ◽  
Colloidal Gold ◽  
Actin Filaments ◽  
Secretory Granules ◽  
Secretory Cells ◽  
Ultrastructural Studies ◽  
Transmission Electron ◽  
Granule Membrane ◽  
Ultrathin Sections ◽  
Lowicryl K4m

In secretory cells a dense and complex network of actin filaments is seen in the subplasmalemmal space attached to the cell membrane. During exocytosis this network is undergoing a rearrangement facilitating access of granules to plasma membrane in order that fusion of the membranes can take place. A filamentous network related to secretory granules has been reported, but its structural organization and composition have not been examined, although this network may be important for exocytosis.Samples of peritoneal mast cells were frozen at -70°C and thawed at 4°C in order to rupture the cells in such a gentle way that the granule membrane is still intact. Unruptured and ruptured cells were fixed in 2% paraformaldehyde and 0.075% glutaraldehyde, dehydrated in ethanol. For TEM (transmission electron microscopy) cells were embedded in Lowicryl K4M at -35°C and for SEM (scanning electron microscopy) they were placed on copper blocks, critical point dried and coated. For immunoelectron microscopy ultrathin sections were incubated with monoclonal anti-actin and colloidal gold labelled IgM. Ruptured cells were also placed on cover glasses, prefixed, and incubated with anti-actin and colloidal gold labelled IgM.

Ultrastructural studies on the interaction of haemophilus influenzae with human endothelial cells in vitro

1990 ◽  
Vol 48 (3) ◽  
pp. 636-637
Author(s):  
D.J.P. Ferguson ◽  
M. Virji ◽  
H. Kayhty ◽  
E.R. Moxon
Keyword(s):  
Endothelial Cells ◽  
Haemophilus Influenzae ◽  
Intercellular Junctions ◽  
Blood Stream ◽  
Ultrastructural Studies ◽  
Endothelial Barriers ◽  
Serotype B ◽  
Meningitis In Children ◽  
Respiratory Epithelial

Haemophilus influenzae is a human pathogen which causes meningitis in children. Systemic H. influenzae infection is largely confined to encapsulated serotype b organisms and is a major cause of meningitis in the U.K. and elsewhere. However, the pathogenesis of the disease is still poorly understood. Studies in the infant rat model, in which intranasal challenge results in bacteraemia, have shown that H. influenzae enters submucosal tissues and disseminates to the blood stream within minutes. The rapidity of these events suggests that H. influenzae penetrates both respiratory epithelial and endothelial barriers with great efficiency. It is not known whether the bacteria penetrate via the intercellular junctions, are translocated within the cells or carried across the cellular barrier in 'trojan horse' fashion within phagocytes. In the present studies, we have challenged cultured human umbilical cord_vein endothelial cells (HUVECs) with both capsulated (b+) and capsule-deficient (b-) isogenic variants of one strain of H. influenzae in order to investigate the interaction between the bacteria and HUVEC and the effect of the capsule.

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