Chloride transport across the membrane of parotid secretory granules

1990 ◽  
Vol 259 (3) ◽  
pp. C413-C420 ◽  
Author(s):  
K. W. Gasser ◽  
U. Hopfer
Keyword(s):  
Chloride Transport ◽  
Secretory Granules ◽  
Disulfonic Acid ◽  
Parotid Glands ◽  
Transport Pathways ◽  
Anion Selectivity ◽  
Granule Membrane ◽  
Cytoplasmic Side ◽  
Cl Conductance

The Cl- transport pathways in secretory granules isolated from the parotid glands of rats were characterized by the technique of ionophore-induced lysis in defined salt solutions. The granules were shown to possess a Cl- conductance that exhibited a distinct anion selectivity with a sequence I- greater than Br- greater than Cl- greater than F- greater than SO4(2-) much greater than gluconate-. This conductance could be reduced approximately 40% by the stilbene 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS) from the cytoplasmic side; the half-maximal concentration for inhibition was 50 microM. Furthermore, the apparent Cl- conductance was reduced by outwardly directed granule H+ gradients and stimulated by inwardly directed gradients. An outwardly directed H+ gradient mimics the in vivo environment and may serve in a regulatory capacity, providing for a tonic inhibition of transport until the granule fuses with the luminal membrane. The granules also possessed a Cl(-)-HCO3- exchange based on electroneutrality of Cl- uptake and stimulation of this uptake by HCO3-. This pathway displayed a different anion selectivity, I- greater than Br- greater than F- greater than Cl- much greater than SO4(2-) much greater than gluconate-, and was not inhibited by SITS on the cytoplasmic side. The presence of these electrolyte transport pathways in the granule membrane is consistent with the production of primary fluid by parotid acinar cells after fusion of granules with the luminal plasma membrane.

The adrenal paraneurone: tubulin organization

1984 ◽  
Vol 62 (5) ◽  
pp. 502-511 ◽  
Author(s):  
M. F. Bader ◽  
F. Bernier-Valentin ◽  
B. Rousset ◽  
D. Aunis
Keyword(s):  
Cell Body ◽  
Intracellular Transport ◽  
Cultured Cells ◽  
Specific Binding ◽  
Secretory Granules ◽  
Immunofluorescent Staining ◽  
Chromaffin Granules ◽  
Two Dimensional Gel Electrophoresis ◽  
Granule Membrane

When chromaffin cells from the bovine adrenal medulla are maintained in culture, they develop neuritelike processes which end with growth-cone-like structures. Chromaffin granules were found to migrate from the cell body to the neurite endings. Thus, the intracellular transport of secretory granules, existing in vivo, seems to occur in an exaggerated way in the cultured cells. These cells offer an excellent model for studying the mechanism of transport, particularly the role of microtubules. By immunofluorescent staining, we observed that tubulin antibodies decorate a complex network visible along the neurites. Colchicine treatment induced the disappearance of this network followed by a return of granules in the cell body and a retraction of neurites. To test the presence of tubulin in the chromaffin granule membrane, we used two-dimensional gel electrophoresis and a radioimmunoassay. Our results indicate that tubulin is not a significant component of chromaffin granules. However, binding experiments show that granule membranes are able to bind tubulin through high affinity binding sites. These results show that microtubules appear involved in neurite formation and probably in granule transport. Tubulin is not an integral constituent of the granule membrane, but is present as a result of a reversible specific binding. This insertion of tubulin into the membrane might represent a step in the association between microtubules and secretory granules.

Intracellular Elemental Concentrations in Resting and Secreting Rat Parotid Glands

1987 ◽  
Vol 66 (2) ◽  
pp. 537-540 ◽  
Author(s):  
K.T. Izutsu ◽  
D.E. Johnson ◽  
M. Goddard
Keyword(s):  
Secretory Granules ◽  
Dry Weight ◽  
Parotid Glands ◽  
X Ray ◽  
Elemental Concentrations ◽  
Saliva Secretion ◽  
Concentration Changes ◽  
Rat Parotid ◽  
Micro Analysis

Electron probe x-ray micro-analysis was used to study the elemental concentration changes that occur during pilocarpine-stimulated saliva secretion. Quantitative x-ray micro-analysis of elemental concentrations in intracellular compartments of rat parotid glands stimulated in vivo with pilocarpine showed that Na concentration was significantly increased, while K concentration was significantly reduced. The magnitude of these changes was consistent with values obtained in other tissues with the x-ray micro-analysis method, and in the same tissue with other experimental methods. Comparisons with results from studies utilizing dispersed acini suggest that acinar dispersion procedures may affect intracellular elemental concentrations. Total electrolyte concentrations in cytoplasm and secretory granules were estimated to increase on a dry-weight basis following pilocarpine stimulation. The former change is consistent with the notion of a trans-cellular route of salivary fluid flow, while the latter change may be important in the exocytosis of secretory granules.

scholarly journals CONCOMITANT SYNTHESIS OF MEMBRANE PROTEIN AND EXPORTABLE PROTEIN OF THE SECRETORY GRANULE IN RAT PAROTID GLAND

1971 ◽  
Vol 50 (1) ◽  
pp. 187-200 ◽  
Author(s):  
Abraham Amsterdam ◽  
Michael Schramm ◽  
Itzhak Ohad ◽  
Yoram Salomon ◽  
Zvi Selinger
Keyword(s):  
Amino Acids ◽  
Secretory Granule ◽  
De Novo ◽  
Secretory Granules ◽  
Specific Radioactivity ◽  
Staining Intensity ◽  
Granule Membrane ◽  
Rat Parotid ◽  
Cell Fractions

After enzyme secretion the membrane of the secretory granule, which had been fused to the cell membrane, was resorbed into the cell. Experiments were therefore carried out to test whether formation of new secretory granules involves reutilization of the resorbed membrane or synthesis of a new membrane, de novo, from amino acids. Incorporation of amino acids-14C into proteins of various cell fractions was measured in vivo, 30, 120, and. 300 min after labeling. At all times the specific radioactivity of the secretory granule membrane was about equal to that of the granule's exportable content. At 120 and 300 min the specific radioactivity of the granule membrane and of the granule content was much higher than that of any other subcellular fraction. It is therefore concluded that the protein of the membrane is synthesized de novo concomitantly with the exportable protein. The proteins of the granule membrane could be distinguished from those of the granule content by gel electrophoresis. All major bands were labeled proportionately to their staining intensity. The amino acid composition of the secretory granule membrane was markedly different from that of the granule's content and also from that of the mitochondrial membrane. The granule membrane showed a high proline content, 30 moles/100 moles amino acids. The analyses show that the radioactivity of the granule membrane is indeed inherent in its proteins and is not due to contamination by other fractions. The possibility is considered that the exportable protein leaves the endoplasmic reticulum already enveloped by the newly synthesized membrane.

Ion transport in rat proximal colon in vivo

1986 ◽  
Vol 251 (1) ◽  
pp. G132-G139 ◽  
Author(s):  
R. Lubcke ◽  
K. Haag ◽  
E. Berger ◽  
H. Knauf ◽  
W. Gerok
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Proximal Colon ◽  
Good Evidence ◽  
Transport Processes ◽  
Disulfonic Acid ◽  
Transport Pathways ◽  
Nacl Absorption ◽  
Ion Substitution

Active Na+ absorption by the rat proximal colon in vivo is for the most part electrically silent. The rheogenic Na+ flux makes up only 8%. To elucidate the underlying transport pathways, the following experimental approaches were used: ion substitution experiments such as choline for Na+, cyclamate for Cl-, variation of luminal pH; administration of known inhibitors; and determination of changes in luminal CO2 tension and pH. The transcolonic ion fluxes as well as the electrical parameters potential difference, specific electrical resistance, and short-circuit current were monitored. Na+ transport was drastically reduced in the absence of luminal Cl-, and vice versa Cl- absorption was blocked at zero Na+. NaCl absorption was blocked by amiloride (10(-3) M) and 4-acetamido-4'-isothiocyanostilbene-2, 2'-disulfonic acid and was lowered by acetazolamide. Colonic NaCl absorption was not influenced by luminal furosemide. Na+ absorption increased with alkalinization of the luminal fluid. Tris instead of HCO-3 buffer at constant pH favored Cl- uptake. The results may easily be explained by the operation of a Na+-H+ antiport functionally coupled to a Cl(-)-HCO-3 antiport. These transport processes are supposed to be present in the columnar cells of the colonic epithelium. There is good evidence for the association of K+ secretion with rheogenic Cl- secretion by the crypt cells.

scholarly journals Inhibitory effect of colchicines on amylase secretion by rat parotid glands: possible localization in the golgi area

1977 ◽  
Vol 73 (3) ◽  
pp. 578-593 ◽  
Author(s):  
C Patzelt ◽  
D Brown ◽  
B Jeanrenaud
Keyword(s):  
De Novo ◽  
Secretory Granules ◽  
Inhibitory Effect ◽  
Binding Activity ◽  
Amylase Secretion ◽  
Parotid Glands ◽  
Secretory Material ◽  
Delayed Effect ◽  
Rat Parotid

Colchicine inhibited amylase secretion by isolated rat parotid glands only 6 h after administration of the drug in vivo. This delayed effect was not the result of the inability of the drug to reach its reaction site. When parotid glands were emptied of their secretory granules by isoproterenol treatment, the subsequent replenishment of cells with granules was inhibited by colchicines. Colchicine concomitantly produced alterations of the Golgi complexes, the cisternae of which were reduced in size and surrounded by clusters of microvesicles. Incubation of parotid glands with colchicines for prolonged durations failed to alter stored amylase secretion as stimulated by isoproterenol, but it inhibited the release of de novo synthesized enzyme. Another colchicines-binding activity, firmly bound to the particular fraction of homogenates, was found, of which a part may represent membrane located microtubular protein. An assembly-disassembly cycle of microtubules appears to exist in the parotid gland, as in the liver. However, only 14 percent of tubulin was found to be polymerized as microtubules in parotid glands as opposed to 40 percent in the liver. The present data suggest that colchicine primarily inhibits the transfer of secretory material towards or away from the Golgi complexes but not the hormone-stimulated secretion of stored amylase.

ATP-sensitive potassium transport by pancreatic secretory granule membrane

1993 ◽  
Vol 264 (1) ◽  
pp. G137-G142
Author(s):  
K. W. Gasser ◽  
J. R. Holda
Keyword(s):  
Secretory Granules ◽  
Pancreatic Acinar Cells ◽  
Dependent Manner ◽  
Transport Pathways ◽  
Comparable Effect ◽  
Granule Membrane ◽  
Fluid Production ◽  
Decreasing Ph ◽  
Dose Dependent ◽  
K Transport

Electrolyte transport pathways in the pancreatic secretory granules may contribute to acini fluid production after fusion with the apical membrane. A component of this granule transport is a K(+)-selective pathway that has been measured indirectly by ionophore-induced lysis of the isolated secretory granules when suspended in a KCl solution. This granule membrane K+ transport was shown to be inhibited by physiological levels of ATP in a dose-dependent manner and was not reversed by ADP. The sulfonylurea tolbutamide (0.5 mM), a recognized inhibitor of ATP-sensitive K+ channels, also reduced the ionophore-dependent lysis by 46%. The ATP sensitivity of the K+ transport was influenced by pH (increased ATP sensitivity with decreasing pH) and KCl concentration (increased ATP sensitivity with increasing KCl). In addition, preincubation with phospholipase A2 (8.3 x 10(-10) g/ml) or lysophospholipids (6.7 x 10(-7) g/ml) produced a significant decrease in the granule K+ transport. However, it is not likely that this inhibition is due to a change in membrane fluidity, because fluidization with arachidonic acid or octanol did not have a comparable effect. The results then support a granule-associated K+ transport in pancreatic acinar cells and suggest that it is ATP and lysophospholipid sensitive.

scholarly journals 5-Hydroxytryptamine transport in cells and secretory granules from a transplantable rat insulinoma

1983 ◽  
Vol 210 (3) ◽  
pp. 803-810 ◽  
Author(s):  
J C Hutton ◽  
M Peshavaria ◽  
N E Tooke
Keyword(s):  
B Cell ◽  
Secretory Granules ◽  
Close Correlation ◽  
Alkaline Media ◽  
Parent Molecule ◽  
Simple Diffusion ◽  
Wide Range ◽  
Granule Membrane ◽  
Rat Insulinoma

Mechanisms of transport of 5-hydroxytryptamine in the pancreatic B-cell were investigated by using cell suspensions and secretory granules prepared from a transplantable rat insulinoma. (1) Cells incubated with 5-hydroxy[G-3H]tryptamine at concentrations ranging from 0.1 microM to 5 mM accumulated the radioisotope principally by a simple diffusion process. The incorporated radioactivity was recovered principally as the parent molecule and was recovered predominantly in soluble protein and secretory-granule fractions prepared from the tissue. (2) Isolated granules incubated in buffered iso-osmotic medium without ATP accumulated the amine to concentrations up to 38-fold that of the medium. This process was insensitive to reserpine and occurred over a wide range of 5-hydroxytryptamine concentrations (0.075 microM-25 mM). Above 5 mM, 5-hydroxytryptamine accumulation decreased in parallel with the breakdown of the delta pH across the granule membrane. Uptake was favoured by alkaline media and was reduced by the addition of (NH4)2SO4. In both cases a close correlation was observed between uptake and the transmembrane delta pH, a finding that suggested that 5-hydroxytryptamine permeated the membrane as the free base and equilibrated across the membrane with the delta pH. Binding of 5-hydroxytryptamine to granule constituents also played a part in this process. ATP caused a further doubling of granule 5-hydroxytryptamine uptake by a process that was sensitive to reserpine (0.5 microM). Inhibitor studies suggested that amine transport in this instance was linked to the activity of the granule membrane proton-translocating ATPase. (3) It was concluded that the uptake of amines driven by proton gradients across the insulin-granule membrane could account for the accumulation in vivo of amines in the B-cell.

Differential aggregation properties of secretory proteins that are stored in exocrine secretory granules of the pancreas and parotid glands

2004 ◽  
Vol 286 (2) ◽  
pp. C365-C371 ◽  
Author(s):  
S. G. Venkatesh ◽  
Darrin J Cowley ◽  
Sven-Ulrik Gorr
Keyword(s):  
Secretory Granules ◽  
Secretory Protein ◽  
Low Ph ◽  
Secretory Proteins ◽  
Parotid Glands ◽  
Granule Membrane ◽  
Parotid Secretory Protein ◽  
Rat Parotid ◽  
Ph Range ◽  
Induced Aggregation

Low-pH- and calcium-induced aggregation of regulated secretory proteins has been proposed to play a role in their retention and storage in secretory granules. However, this has not been tested for secretory proteins that are stored in the exocrine parotid secretory granules. Parotid granule matrix proteins were analyzed for aggregation in the presence or absence of calcium and in the pH range of 5.5 to 7.5. Amylase did not aggregate under these conditions, although <10% of parotid secretory protein (PSP) aggregated below pH 6.0. To test aggregation directly in isolated granules, rat parotid secretory granules were permeabilized with 0.1% saponin in the presence or absence of calcium and in the pH range of 5.0 to 8.4. In contrast to the low-pH-dependent retention of amylase in exocrine pancreatic granules, amylase was quantitatively released and most PSP was released from parotid granules under all conditions. Both proteins were completely released upon granule membrane solubilization. Thus neither amylase nor PSP show low-pH- or calcium-induced aggregation under physiological conditions in the exocrine parotid secretory granules.

Thiazide-sensitive sodium chloride cotransport in early distal tubule

1987 ◽  
Vol 253 (3) ◽  
pp. F546-F554 ◽  
Author(s):  
D. H. Ellison ◽  
H. Velazquez ◽  
F. S. Wright
Keyword(s):  
Sodium Transport ◽  
Chloride Transport ◽  
Collecting Duct ◽  
Distal Tubule ◽  
Sodium Absorption ◽  
Transport Pathways ◽  
Luminal Membrane ◽  
Chloride Absorption ◽  
Distal Tubules

At least two pathways mediate sodium absorption across the luminal membrane of the renal distal tubule. One pathway is a conductive channel and the other appears to be a coupled Na-Cl cotransport pathway. The distal tubule comprises three segments: the distal convoluted tubule, the connecting tubule, and the initial collecting duct. To provide information about cellular locations of the proposed sodium transport pathways, we perfused early (14-38% of whole distal length) and late (61-83% of whole distal length) segments of whole distal tubules separately in vivo in anesthetized rats. When perfused with a solution that resembles fluid normally arriving at the distal tubule (75 mM Na, 68 mM Cl), rates of sodium absorption were similar in early and late segments (early 68 +/- 29.6, late 67 +/- 27.5 pmol X min-1 X mm-1). When perfused with a solution that resembles interstitial fluid (148 mM Na, 110 mM Cl), sodium transport was significantly higher in early than in late segments (276 +/- 28.4 vs. 113 +/- 29.7 pmol X min-1 X mm-1). Chlorothiazide (10(-3) M), which blocks sodium and chloride absorption in whole distal tubules, reduced sodium and chloride transport to zero in early distal tubules but had no significant effect in late distal tubules. Removing all chloride from perfusion solutions reduced sodium transport in early but not late distal segments.(ABSTRACT TRUNCATED AT 250 WORDS)

Chloride transport across the basolateral membrane of rabbit proximal convoluted tubules

1990 ◽  
Vol 258 (6) ◽  
pp. F1569-F1578 ◽  
Author(s):  
K. Ishibashi ◽  
F. C. Rector ◽  
C. A. Berry
Keyword(s):  
Chloride Transport ◽  
Basolateral Membrane ◽  
Disulfonic Acid ◽  
Transport Mechanisms ◽  
Kcl Cotransport ◽  
Cl Transport ◽  
Convoluted Tubules ◽  
Cl Conductance ◽  
Proximal Convoluted Tubules

To examine the basolateral Cl transport mechanisms of proximal convoluted tubules (PCT), intracellular Cl activity (AiCl) was measured with double-barreled Cl-selective microelectrodes. When rabbit PCT were perfused in vitro with high Cl, low HCO3, and bathed with ultrafiltrate-like solutions, AiCl was 29.9 +/- 0.4 mM and basolateral membrane voltage (Vbl) was -47.7 +/- 0.4 mV (n = 247). Possible basolateral Cl transport mechanisms that we examined were as follows: Cl conductance, KCl cotransport, and Na-dependent Cl-HCO3 exchange. Cl conductance was negligible, since the voltage clamp of Vbl to 30 mV above and below the spontaneous Vbl did not change AiCl even in the absence of luminal Cl. KCl cotransport was suggested by 1) increasing bath K, increased AiCl, and 2) decreasing bath K decreased AiCl. KCl cotransport was Na independent and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS), barium, and furosemide insensitive. Na-dependent Cl-HCO3 exchange was suggested by 1) bath HCO3 reduction increased AiCl, which was greatly inhibited by bath Na removal or bath SITS, and 2) bath Na removal increased AiCl, which was completely blocked by bath SITS. We conclude that 1) Cl conductance is negligibly small at the basolateral membrane and 2) SITS-insensitive KCl cotransport and SITS-sensitive Na-dependent Cl-HCO3 exchange are present at the basolateral membrane.

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