scholarly journals Ultrastructural Organization of Bovine Chromaffin Cell Cortex—Analysis by Cryofixation and Morphometry of Aspects Pertinent to Exocytosis

1997 ◽  
Vol 139 (7) ◽  
pp. 1709-1717 ◽  
Author(s):  
Helmut Plattner ◽  
Antonio R. Artalejo ◽  
Erwin Neher
Keyword(s):  
Average Diameter ◽  
Immediate Release ◽  
Cell Cortex ◽  
Ultrastructural Organization ◽  
Potential Contribution ◽  
High K ◽  
Area Increase ◽  
Ultrathin Sections ◽  
Bovine Chromaffin Cells ◽  
Measured Size Distribution

We have analyzed ultrathin sections from isolated bovine chromaffin cells grown on plastic support, after fast freezing, by quantitative electron microscopy. We determined the size and intracellular distribution of dense core vesicles (DVs or chromaffin granules) and of clear vesicles (CVs). The average diameter of DVs is 356 nm, and that of CVs varies between 35–195 nm (average 90 nm). DVs appear randomly packed inside cells. When the distance of the center of DVs to the cell membrane (CM) is analyzed, DV density is found to decrease as the CM is approached. According to Monte Carlo simulations performed on the basis of the measured size distribution of DVs, this decay can be assigned to a “wall effect.” Any cortical barrier, regardless of its function, seems to not impose a restriction to a random cortical DV packing pattern. The number of DVs closely approaching the CM (docked DVs) is estimated to be between 364 and 629 (average 496), i.e., 0.45 to 0.78 DVs/μm2 CM. Deprivation of Ca2+, priming by increasing [Ca2+]i, or depolarization by high [K+]e for 10 s (the effect of which was controlled electrophysiologically and predicted to change the number of readily releasable granules [RRGs]) does not significantly change the number of peripheral DVs. The reason may be that (a) structural docking implies only in part functional docking (capability of immediate release), and (b) exocytosis is rapidly followed by endocytosis and replenishment of the pool of docked DVs. Whereas the potential contribution of DVs to CM area increase by immediate release can be estimated at 19–33%, that of CVs is expected to be in the range of 5.6–8.0%.

Different contributions of L- and Q-type Ca2+ channels to Ca2+ signals and secretion in chromaffin cell subtypes

1997 ◽  
Vol 272 (2) ◽  
pp. C476-C484 ◽  
Author(s):  
R. B. Lomax ◽  
P. Michelena ◽  
L. Nunez ◽  
J. Garcia-Sancho ◽  
A. G. Garcia ◽  
...  
Keyword(s):  
Chromaffin Cells ◽  
Chromaffin Cell ◽  
Channel Blocker ◽  
Cell Types ◽  
Bay K 8644 ◽  
High K ◽  
Voltage Dependent ◽  
Bovine Chromaffin Cells ◽  
P Type ◽  
Ca2 Channels

In this study, we investigated the contribution of different subtypes of voltage-dependent Ca2+ channels to changes in cytosolic free Ca2+ ([Ca2+]i) and secretion in noradrenergic and adrenergic bovine chromaffin cells. In single immunocytochemically identified chromaffin cells, [Ca2+]i increased transiently during high K+ depolarization. Furnidipine and BAY K 8644, L-type Ca2+ channel blocker and activator, respectively, affected the [Ca2+]i rise more in noradrenergic than in adrenergic cells. In contrast, the Q-type Ca2+ channel blocker omega-conotoxin MVIIC inhibited the [Ca2+]i rise more in adrenergic cells. omega-Agatoxin IVA (30 nM), which blocks P-type Ca2+ channels, had little effect on the [Ca2+]i signal. The N-type Ca2+ channel blocker omega-conotoxin GVIA similarly inhibited the [Ca2+]i rise in both cell types. The effects of furnidipine, BAY K 8644, and omega-conotoxin MVIIC on K+-evoked norepinephrine and epinephrine release paralleled those effects on [Ca2+]i signals. However, omega-conotoxin GVIA and 30 nM omega-agatoxin IVA did not affect the secretion of either amine. The data suggest that, in the bovine adrenal medulla, the release of epinephrine and norepinephrine are preferentially controlled by Q- and L-type Ca2+ channels, respectively. P- and N-type Ca2+ channels do not seem to control the secretion of either catecholamine.

scholarly journals Insulin unmasks a COOH-terminal Glut4 epitope and increases glucose transport across T-tubules in skeletal muscle.

1996 ◽  
Vol 135 (2) ◽  
pp. 415-430 ◽  
Author(s):  
W Wang ◽  
P A Hansen ◽  
B A Marshall ◽  
J O Holloszy ◽  
M Mueckler
Keyword(s):  
Skeletal Muscle ◽  
Glucose Transport ◽  
Insulin Treatment ◽  
Average Diameter ◽  
Membrane Domains ◽  
Transverse Tubules ◽  
Confocal Immunofluorescence Microscopy ◽  
Confocal Immunofluorescence ◽  
Ultrathin Sections

An improved immunogold labeling procedure was used to examine the subcellular distribution of glucose transporters in Lowricryl HM20-embedded skeletal muscle from transgenic mice overexpressing either Glut1 or Glut4. In basal muscle, Glut4 was highly enriched in membranes of the transverse tubules and the terminal cisternae of the triadic junctions. Less than 10% of total muscle Glut4 was present in the vicinity of the sarcolemmal membrane. Insulin treatment increased the number of gold particles associated with the transverse tubules and the sarcolemma by three-fold. However, insulin also increased the total Glut4 immunogold reactivity in muscle ultrathin sections by up to 1.8-fold and dramatically increased the amount of Glut4 in muscle sections as observed by laser confocal immunofluorescence microscopy. The average diameter of transverse tubules observed in longitudinal sections increased by 50% after insulin treatment. Glut1 was highly enriched in the sarcolemma, both in the basal state and after insulin treatment. Disruption of transverse tubule morphology by in vitro glycerol shock completely abolished insulin-stimulated glucose transport in isolated rat epitrochlearis muscles. These data indicate that: (a) Glut1 and Glut4 are targeted to distinct plasma membrane domains in skeletal muscle; (b) Glut1 contributes to basal transport at the sarcolemma and the bulk of insulin-stimulated transport is mediated by Glut4 localized in the transverse tubules; (c) insulin increases the apparent surface area of transverse tubules in skeletal muscle; and (d) insulin causes the unmasking of a COOH-terminal antigenic epitope in skeletal muscle in much the same fashion as it does in rat adipocytes.

Plasmalemmal sodium-calcium exchanger shapes the calcium and exocytotic signals of chromaffin cells at physiological temperature

2013 ◽  
Vol 305 (2) ◽  
pp. C160-C172 ◽  
Author(s):  
Juan-Fernando Padín ◽  
José-Carlos Fernández-Morales ◽  
Román Olivares ◽  
Stefan Vestring ◽  
Juan-Alberto Arranz-Tagarro ◽  
...  
Keyword(s):  
Chromaffin Cells ◽  
Catecholamine Release ◽  
Sodium Calcium Exchanger ◽  
Physiological Temperature ◽  
Slowing Down ◽  
Sodium Calcium ◽  
High K ◽  
Highly Sensitive ◽  
Bovine Chromaffin Cells ◽  
Kinetics Of

The activity of the plasmalemmal Na+/Ca2+ exchanger (NCX) is highly sensitive to temperature. We took advantage of this fact to explore here the effects of the NCX blocker KB-R7943 (KBR) at 22 and 37°C on the kinetics of Ca2+ currents ( ICa), cytosolic Ca2+ ([Ca2+]c) transients, and catecholamine release from bovine chromaffin cells (BCCs) stimulated with high K+, caffeine, or histamine. At 22°C, the effects of KBR on those parameters were meager or nil. However, at 37°C whereby the NCX is moving Ca2+ at a rate fivefold higher than at 22°C, various of the effects of KBR were pronounced, namely: 1) no effects on ICa; 2) reduction of the [Ca2+]c transient amplitude and slowing down of its rate of clearance; 3) blockade of the K+-elicited quantal release of catecholamine; 4) blockade of burst catecholamine release elicited by K+; 5) no effect on catecholamine release elicited by short K+ pulses (1–2 s) and blockade of the responses produced by longer K+ pulses (3–5 s); and 6) potentiation of secretion elicited by histamine or caffeine. Furthermore, the more selective NCX blocker SEA0400 also potentiated the secretory responses to caffeine. The results suggest that at physiological temperature the NCX substantially contributes to shaping the kinetics of [Ca2+]c transients and the exocytotic responses elicited by Ca2+ entry through Ca2+ channels as well as by Ca2+ release from the endoplasmic reticulum.

Effect of depolarization by potassium proprionate on synaptic vesicle diameter and distribution of the snake motor endplate

1989 ◽  
Vol 47 ◽  
pp. 946-947
Author(s):  
Gregory Hendricks ◽  
Lisa Coniglio ◽  
Ian Marshall ◽  
Tim Searl ◽  
Rodney Parsons
Keyword(s):  
Synaptic Vesicles ◽  
Neuromuscular Junctions ◽  
Muscle Fibers ◽  
Average Diameter ◽  
Uranyl Acetate ◽  
Release Agent ◽  
High K ◽  
Microscope Slides ◽  
Whole Mounts

The morphological effects of elevated potassium (K+) at the neuromuscular junction of garter snake twitch muscle fibers were examined in vitro. Elevated K+ depolarizes the motor neuron terminals and causes quantal release of acetylcholine. It has been shown in amphibians that prolonged (30-60 min) depolarization with high K+ depletes terminals of synaptic vesicles ; but causes no change in the average diameter of the vesicles. In the snake, however, we have shown previously that miniature endplate currents (MEPCs) are present at high frequency even after 1-2 hours of exposure to high K+ . This indicates that in the snake, synaptic vesicles are not totally depleted by depolarization with high K+. In this study, we examined the effect of depolarization with high K+ on the distribution and diameter of synaptic vesicles.Intercostal muscle preparations were exposed for 15 minutes to Ringer's + solution containing either a control (1.2mM) or high (70mM) concentration of K+ Preparations were fixed immediately in 1% glutaraldehyde for 15 minutes and washed with Millonig's buffer. The whole mount muscle preparations were then fixed for 1 h in 2.5% glutaraldehyde in 0.1 M phosphate buffer, pH 7.2; rinsed three times in fresh buffer and post-fixed in 2% OsO4 in the same buffer for 30 min, rinsed in fresh buffer and dehydrated in a graded ethanol series to 100% and then transferred into a 2% solution of uranyl acetate for 20 min to en-block stain the whole mounts. The preps were then transferred back to 100% ethanol to complete the dehydration and embedded in a mixture of Embed 812/Araldite 502 (Hard) between two microscope slides that had been precoated with Formen- Trennmittel Liquid Release Agent and polymerized overnight at 70°C.The whole mounts were then removed from the microscope slides and examined on a compound light microscope where the neuromuscular junctions on the twitch muscle fibers could be identified, marked and cut out of the preps. These pieces of the whole mounts were then remounted into a slot on the end of a pre-cast Embed-Araldite block with a drop of fresh resin and polymerized.

Fluid-phase uptake by macropinocytosis in Dictyostelium

1997 ◽  
Vol 110 (2) ◽  
pp. 105-112 ◽  
Author(s):  
U. Hacker ◽  
R. Albrecht ◽  
M. Maniak
Keyword(s):  
Fluorescent Protein ◽  
Fluid Phase ◽  
Average Diameter ◽  
Cell Cortex ◽  
Dictyostelium Cell ◽  
Scanning Electron Micrographs ◽  
Extracellular Medium ◽  
Green Fluorescent ◽  
Cytoskeletal Rearrangements ◽  
Shape Changes

To study fluid-phase endocytosis in living cells and its relationship to changes in the cell cortex, we have used a green fluorescent protein (GFP)-tagged version of coronin, an actin-associated protein that localises to dynamic regions of the Dictyostelium cell cortex. In the confocal microscope, internalisation of fluorescently labelled dextran as a fluid-phase marker can be recorded simultaneously with the recruitment of the coronin-GFP fusion-protein from the cytoplasm of the phagocyte. At crown-shaped surface protrusions, extracellular medium is taken up into vesicles with an average diameter of 1.6 microns, which is significantly larger than the 0.1 microns diameter of clathrin-coated pinosomes. The observed frequency of macropinosome formation can account for a large portion, if not all, of the fluid-phase uptake. The redistribution of coronin-GFP strongly resembles cytoskeletal rearrangements during phagocytosis. Scanning-electron micrographs indicate that crown-shaped cell-surface extensions can undergo shape changes, without a particle bound, that are similar to shape changes that occur during phagocytosis. In quantitative assays, the uptake of particles and fluid are about equally dependent on F-actin and coronin.

scholarly journals ULTRASTRUCTURAL ORGANIZATION OF NEURONS OF THE LATERAL PREOPTIC NUCLEUS OF THE HYPOTHALAMUS OF OLD RATS UNDER VARIOUS LIGHT CONDITIONS

2021 ◽  
Vol 20 (3) ◽  
Author(s):  
V.R. Yosypenko ◽  
R.Ye. Bulyk ◽  
M.I. Kryvchanska ◽  
Y.R. Lukan
Keyword(s):  
Epoxy Resins ◽  
Dark Period ◽  
White Male ◽  
Ultrastructural Organization ◽  
Light Conditions ◽  
Preoptic Nucleus ◽  
Cell Nuclei ◽  
Light Mode ◽  
Old Rats ◽  
Ultrathin Sections

Purpose – to study the changes in the ultrastructure of neurons of the lateral preopticnucleus (LPON) of the hypothalamus of old rats under various light conditions.Material and methods. The experiments were carried out on 36 old white male rats.The material was fixed in a 2.5% solution of glutaraldehyde, prepared on the basisof phosphate buffer with a pH of 7.2–7.4. Then, postfixation was performed in a 1%solution of osmium tetraoxide and dehydrated in propylene oxide, after which it waspoured into a mixture of epoxy resins. Ultrathin sections made on an ultramicrotomeLKB-3 were contrasted with uranium acetate and lead citrate according to the Reynoldsmethod and studied under electron microscope TEM - 125 K.Results. Studies of LPON neurons under the standard light mode revealed nuclei withuneven contours. The nucleoli are quite large. The neuroplasm contains well-developedtubules of the rough endoplasmic reticulum (ER) and small cisternae of the Golgi complex(GC). Mitochondria are rounded, small, with moderately pronounced cristae. Underconditions of round-the-clock darkness, we have found that the cell nuclei are rounded,less often determined by the nucleoli. In the neuroplasm there are locally dilated tubulesof the ER and cisternae of the GC, mitochondria with enlightened matrix and fragmentedcristae. Under conditions of round-the-clock illumination, the nuclei of the rounded formwith uneven contours of a nuclear membrane forming deep intussusception are revealed.Nucleoli were rarely identified. In the hyaloplasm, locally dilated tubules of the ERare identified. Mitochondria are small in size with an enlightened matrix and reducedcristae.Conclusions. The obtained results of submicroscopic examination of LPON neuronsof the hypothalamus of old rats revealed their relatively increased functional activityin the dark period. Under conditions of round-the-clock lighting, more pronouncedhypertrophic and initial destructive changes of the nuclei and organelles of the neuronsof the LPON of the hypothalamic were revealed, compared with the animals that wereunder the conditions of round-the-clock darkness. This is confirmed by the change in theultrastructure of nerve cells at 2 am the appearance of "dark" cells.

Differential Ca2+ responses of adrenergic and noradrenergic chromaffin cells to various secretagogues

1995 ◽  
Vol 269 (6) ◽  
pp. C1540-C1546 ◽  
Author(s):  
L. Nunez ◽  
M. T. De La Fuente ◽  
A. G. Garcia ◽  
J. Garcia-Sancho
Keyword(s):  
Chromaffin Cells ◽  
Cell Types ◽  
Membrane Receptors ◽  
Differential Distribution ◽  
High K ◽  
Voltage Dependent ◽  
Functional Membrane ◽  
Bovine Chromaffin Cells ◽  
Differential Contribution ◽  
Ca2 Channels

The effects of several physiological agonists on the cytosolic Ca2+ concentration ([Ca2+]i) of immunnocytochemically identified single adrenergic and noradrenergic bovine chromaffin cells were compared. No differences were observed in the responses to stimulation by high-K+ solutions with or without BAY K 8644, suggesting that the density and properties of voltage-dependent Ca2+ channels were similar in both cell types. The increase of [Ca2+]i induced by acetylcholine was greater in adrenergic cells, and this was due to differences in the response mediated through nicotinic receptors. The responses to bradykinin and to ATP were slightly greater in noradrenergic cells. Only a small fraction of the cells (18-28%) was responsive to ATP. The responses to angiotensin II and to histamine were much greater in adrenergic than in noradrenergic cells. Histamine was almost a selective stimulator of adrenergic cells. These differences suggest differential distribution of functional membrane receptors in both cell types and may be relevant to understanding the differential contribution of epinephrine- and norepinephrine-secreting cells during stressful conflicts in physiological or pathophysiological situations.

scholarly journals Directly-Patternable Bi2O3 Nanoparticle for Polymer Nanocomposite Capacitor

Coatings ◽  
2020 ◽  
Vol 10 (8) ◽  
pp. 752
Author(s):  
Ki-Hoon Son ◽  
Hong-Sub Lee
Keyword(s):  
Uv Irradiation ◽  
Nanocomposite Film ◽  
Polyvinylidene Fluoride ◽  
Polymer Nanocomposite ◽  
Average Diameter ◽  
Pvdf Film ◽  
High K ◽  
Mask Aligner ◽  
Metal Organic ◽  
Bi Nanoparticles

A polyvinylidene fluoride (PVDF) film incorporating size-controlled, uniformly dispersed, directly patterned Bi2O3 nanoparticles was developed to achieve a high-k polymer nanocomposite capacitor. The photochemical metal-organic deposition (PMOD) method was employed to form uniformly dispersed and directly patterned nanoparticles on the substrate. Bi nanoparticles were produced by spin coating a Bismuth 2-ethylhexanoate solution on a Pt substrate with UV irradiation for 1, 4, 7, and 10 min. The average diameter of nanoparticles and the number of nanoparticles per unit area (μm2) were about 30, 70, and 120 nm and 30, 30, and 31 particles/μm2 for UV irradiation times of 4, 7, and 10 min, respectively. In addition, the capacitance of PVDF nanocomposite film could be controlled by the Bi2O3 nanoparticle size. The PVDF nanocomposite film containing Bi2O3 nanoparticles with 1, 4, 7, and 10 min UV irradiation were able to improve capacitance by about 1.4, 2.0, 2.7, and 3.4 times compared with an as-prepared PVDF film. By using a mask aligner, directly pattered Bi nanoparticles on the substrate, which had a 5 μm line width pattern, were successfully defined and demonstrated.

Phosphore, calcium, silicium et fer dans des cristaux, extraits de graines sèches de Raphanus

1984 ◽  
Vol 62 (11) ◽  
pp. 2394-2402 ◽  
Author(s):  
Louis Genevès ◽  
Jacques Rutin ◽  
Sylvain Halpern
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Electron Beam ◽  
Electron Probe ◽  
Average Diameter ◽  
Transmission Electron ◽  
Polymorphic Crystals ◽  
Parenchyma Cells ◽  
Ultrathin Sections ◽  
Wavelength Dispersive Spectrometer

Samples were taken from dry seeds of radish and fixed in glutaraldehyde. Ultrathin sections were observed without contrasting treatment. From cotyledonary parenchyma, it was possible to obtain powders or prints, which were observed by transmission electron microscopy. They showed several types of crystals. In the ultrathin sections of parenchyma cells, the crystals are included in globoids. Electron probe microanalysis with a wavelength dispersive spectrometer (Camebax microprobe) showed that they were rich in P, Ca, and Mg. In the powders and the prints, several polymorphic crystals, of varied sizes, were observed; these were sensitive to the electron beam. Some have relatively high ratios in Ca, lower ratios of S, and other elements, such as Si. Others possessed high ratios of Si with other elements, such as Ca and Al. The latter were less dense, more stable under the beam and their average diameter was smaller. Other crystals were smaller (some tenths of a micrometre). They were electron dense and very stable. Some of these were rich in Fe and could contain other elements (among others Si, Ca and P).

Permeation and inactivation by calcium and manganese of bovine adrenal chromaffin cell calcium channels

1992 ◽  
Vol 263 (4) ◽  
pp. C818-C824 ◽  
Author(s):  
R. I. Fonteriz ◽  
J. Garcia-Sancho ◽  
L. Gandia ◽  
M. G. Lopez ◽  
A. G. Garcia
Keyword(s):  
Chromaffin Cells ◽  
Chromaffin Cell ◽  
Divalent Cations ◽  
Receptor Activation ◽  
Adrenal Chromaffin Cell ◽  
High K ◽  
Bovine Chromaffin Cells ◽  
Adrenal Chromaffin ◽  
Ca2 Channels ◽  
Stimulation Of

Stimulation of fura-2-loaded bovine chromaffin cells with the nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP; 10 microM) or depolarization with high [K+] (50 mM) accelerated the entry of both Ca2+ and Mn2+, used here as a Ca2+ surrogate for Ca2+ channels. Removal of extracellular Na+ prevented the effects of DMPP but did not modify the effects of K+, indicating that Na+ is necessary for coupling of Ca2+ entry to the nicotinic receptor activation and that the ionophore associated with it is functionally impermeable to divalent cations. DMPP- as well as K(+)-evoked Ca2+ and Mn2+ influx were blocked completely by Ni2+ but only partially by dihydropyridines, suggesting that, in addition to L-type Ca2+ channels, other Ca2+ entry pathways may be present. Inactivation of Ca2+ channels, followed by comparing the rates of Mn2+ uptake at different time periods after the addition of DMPP or high K+, did not happen in the absence of extracellular Ca2+. When 1 mM Ca2+ was present, a delayed inhibition (half time, 10-20 s) was observed, suggesting that it is not due to the entry of Ca2+ itself but to the increase of the cytoplasmic Ca2+ concentration ([Ca2+]i) that takes a few seconds to develop. The influx of Ca2+, estimated from the increase of [Ca2+]i, was also impaired in a time-dependent fashion by previous entry of Mn2+. Inactivation of Ca2+ entry was achieved at estimated mean intracellular Mn2+ concentrations as low as 10(-9) M.

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