scholarly journals Activation of alpha-2-adrenoceptors results in an increase in F-actin formation in HIT-T15 pancreatic B-cells

1995 ◽  
Vol 307 (1) ◽  
pp. 169-174 ◽  
Author(s):  
H C Cable ◽  
A el-Mansoury ◽  
N G Morgan
Keyword(s):  
Plasma Membrane ◽  
Insulin Secretion ◽  
B Cells ◽  
Fluorescence Microscopy ◽  
Pertussis Toxin ◽  
Actin Polymerization ◽  
Secretory Granules ◽  
Rhodamine Phalloidin ◽  
Late Step ◽  
Pancreatic B Cells

1. Alpha-2-adrenoceptor agonists, such as noradrenaline, are potent inhibitors of insulin secretion, and it has been suggested that they control a late step in the pathway of exocytosis. We have investigated whether this could be related to a change in the extent of actin polymerization in the pancreatic B-cell, since actin microfilaments are implicated in regulating the access of secretory granules to the plasma membrane prior to exocytosis. 2. Cultured HIT-T15 pancreatic B-cells responded to noradrenaline with an increase in F-actin content, as judged by a rise in the fluorescence output after probing of the cells with phalloidin (a toxin which binds specifically to F-actin) conjugated to rhodamine. The response to noradrenaline was rapid, dose-dependent and sustained and could be reproduced by the highly selective alpha-2-agonist UK14,304. Examination of HIT-T15 cells by fluorescence microscopy after treatment with rhodamine-phalloidin, revealed a significant localization of F-actin immediately adjacent to the plasma membrane. The pattern of F-actin distribution in the cells was not altered dramatically by noradrenaline, although the intensity of staining close to the plasma membrane appeared to be slightly reduced. 3. The increase in F-actin content induced by noradrenaline and UK14,304 was inhibited significantly by the alpha-2-antagonist idazoxan but not by the alpha-1-selective antagonist prazosin. Pretreatment of HIT-T15 cells with pertussis toxin did not lead to any direct alteration in F-actin content, although the toxin significantly modified the responses induced by noradrenaline and UK14,304. In each case, cells incubated for 24 h with pertussis toxin responded to the alpha-2-agonist with an enhanced fluorescence output, indicating that F-actin levels had increased still further. This did not correlate with any gross change in the distribution of F-actin as judged by fluorescence microscopy. 4. The results demonstrate that alpha-2-adrenoceptors are coupled to control of actin polymerization in HIT-T15 cells. They suggest that regulation of F-actin formation could be a component of the mechanism by which alpha-2-agonists mediate inhibition of insulin secretion.

scholarly journals The possible mechanisms by which phanoside stimulates insulin secretion from rat islets

2007 ◽  
Vol 192 (2) ◽  
pp. 389-394 ◽  
Author(s):  
Nguyen Khanh Hoa ◽  
Åke Norberg ◽  
Rannar Sillard ◽  
Dao Van Phan ◽  
Nguyen Duy Thuan ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
B Cells ◽  
Pancreatic Islets ◽  
Insulin Release ◽  
Pertussis Toxin ◽  
Insulin Response ◽  
Gk Rat ◽  
Isolated Pancreatic Islets ◽  
Gk Rats ◽  
Rat Islets

We recently showed that phanoside, a gypenoside isolated from the plant Gynostemma pentaphyllum, stimulates insulin secretion from rat pancreatic islets. To study the mechanisms by which phanoside stimulates insulin secretion. Isolated pancreatic islets of normal Wistar (W) rats and spontaneously diabetic Goto-Kakizaki (GK) rats were batch incubated or perifused. At both 3.3 and 16.7 mM glucose, phanoside stimulated insulin secretion several fold in both W and diabetic GK rat islets. In perifusion of W islets, phanoside (75 and 150 μM) dose dependently increased insulin secretion that returned to basal levels when phanoside was omitted. When W rat islets were incubated at 3.3 mM glucose with 150 μM phanoside and 0.25 mM diazoxide to keep K-ATP channels open, insulin secretion was similar to that in islets incubated in 150 μM phanoside alone. At 16.7 mM glucose, phanoside-stimulated insulin secretion was reduced in the presence of 0.25 mM diazoxide (P<0.01). In W islets depolarized by 50 mM KCl and with diazoxide, phanoside stimulated insulin release twofold at 3.3 mM glucose but did not further increase the release at 16.7 mM glucose. When using nimodipine to block L-type Ca2+ channels in B-cells, phanoside-induced insulin secretion was unaffected at 3.3 mM glucose but decreased at 16.7 mM glucose (P<0.01). Pretreatment of islets with pertussis toxin to inhibit exocytotic Ge-protein did not affect insulin response to 150 μM phanoside. Phanoside stimulated insulin secretion from Wand GK rat islets. This effect seems to be exerted distal to K-ATP channels and L-type Ca2+ channels, which is on the exocytotic machinery of the B-cells.

Patterns of calcium localization in pancreatic endocrine cells

1976 ◽  
Vol 21 (1) ◽  
pp. 107-117
Author(s):  
M. Ravazzola ◽  
F. Malaisse-Lagae ◽  
M. Amherdt ◽  
A. Perrelet ◽  
W.J. Malaisse ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
B Cells ◽  
Endocrine Cells ◽  
Secretory Granules ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Ionophore A23187 ◽  
Calcium Localization ◽  
A Cells ◽  
D Cells

Subcellular calcium localization in the dndocrine cells of rat pancreas was studied by the pyroantimonate precipitation technique. Calcium-containing electron-dense deposits in the endocrine cells were mostly found within secretory granules and along the plasma membrane, but their pattern of distribution in A-, B- and D-cells displayed qualitative and quantitative differences. In B-cells, numerous secretory granules contained deposits located in the halo surrounding the granule core. In A-cells, only few granules contained precipitates in their halo, whereas in D-cells, deposits were situated in the dense core of the secretory granules. Deposits along the plasma membrane occurred generally on the outer leaflet of the plasma membrane of B- and D-cells and on the inner leaflet of that of A-cells. In islets incubated at a high glucose concentration or in the presence of the calcium ionophore A23187, the number of beta granules containing precipitates was significantly increased. By contrast, only few deposits were observed in B-cells incubated in calcium-deprived medium enriched with EGTA. These findings indicate that: the pattern of calcium localization varies in different islet cell types; in B-cells the secretory granules represent one of the major stores of intracellular calcium; and that this store undergoes changes in conditions which alter insulin release.

Chronic Lymphocytic Leukemia B Cells Express Functional CXCR4 Chemokine Receptors That Mediate Spontaneous Migration Beneath Bone Marrow Stromal Cells

Blood ◽  
1999 ◽  
Vol 94 (11) ◽  
pp. 3658-3667 ◽  
Author(s):  
Jan A. Burger ◽  
Meike Burger ◽  
Thomas J. Kipps
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Pertussis Toxin ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Actin Polymerization ◽  
Kinase Inhibitor ◽  
Critical Role ◽  
Lymphocytic Leukemia ◽  
Marrow Stromal Cells

Chemokines play a central role for lymphocyte trafficking and homing. The mechanisms that direct the tissue localization of B cells from patients with chronic lymphocytic leukemia (B-CLL) are unknown. We found that CLL B cells express functional CXCR4 receptors for the chemokine stromal cell-derived factor-1 (SDF-1), as demonstrated by receptor endocytosis, calcium mobilization, and actin polymerization assays. Moreover, CLL B cells displayed chemotaxis to this chemokine that could be inhibited by monoclonal antibodies (MoAbs) against CXCR4, pertussis toxin, or Wortmannin, a phosphatidylinositol 3-kinase inhibitor. That this chemotaxis may be involved in the homing of CLL cells is argued by studies in which CLL B cells were cocultured with a murine marrow stromal cell line that secretes SDF-1. Within 2 hours, CLL B cells spontaneously migrated beneath such stromal cells in vitro (pseudoemperipolesis). This migration could be inhibited by pretreatment of CLL B cells with anti-CXCR4 MoAbs, SDF-1, or pertussis-toxin. Furthermore, we noted strong downmodulation of CXCR4 on CLL B cells that migrated into the stromal cell layer. These findings demonstrate that the chemokine receptor CXCR4 on CLL B cells plays a critical role for heterotypic adherence to marrow stromal cells and provide a new mechanism to account for the marrow infiltration by neoplastic B cells.

Glucokinase in pancreatic B-cells and its inhibition by alloxan

1987 ◽  
Vol 115 (1) ◽  
pp. 21-29 ◽  
Author(s):  
Sigurd Lenzen ◽  
Markus Tiedge ◽  
Uwe Panten
Keyword(s):  
Insulin Secretion ◽  
B Cells ◽  
B Cell ◽  
Metabolic Flux ◽  
Inhibitory Concentration ◽  
Pancreatic B Cell ◽  
High K ◽  
Pancreatic B Cells ◽  
Low K

Abstract. Characterization of glucokinase in pancreatic B-cells from ob/ob mice and from rat liver revealed identical characteristics. A narrow substrate specificity; high Km values for the two substrates, D-glucose and D-mannose, in the range of 10 and 20 mmol/l, respectively; higher Vmax values for D-glucose than for D-mannose; inhibition of glucokinase activities by D-mannoheptulose and by a specific glucokinase antibody. These characteristics distinguish glucokinase in soluble cytoplasmic fractions of pancreatic B-cells and liver from low Km hexokinases. Alloxan is a pancreatic B-cell cytotoxic agent, which has been widely used as a tool for the elucidation of the mechanisms of insulin secretion, because its inhibitory action on insulin secretion has been presumed to be intimately related to the mechanism of glucose-induced insulin secretion. Alloxan inhibited glucokinase but not hexokinase activity in cytoplasmic fractions of pancreatic B-cells and liver. The half maximal inhibitory concentration of alloxan was 5 μmol/l. Glucokinase activity was protected from alloxan toxicity only by D-glucose and D-mannose; the α anomer of D-glucose provided significantly greater protection than the β anomer. The non-metabolizable sugar 3-0-methyl-D-glucose did not provide protection of glucokinase activity against inhibition by alloxan. Thus, inhibition of pancreatic B-cell glucokinase may contribute to the inhibition of glucose-induced insulin secretion by alloxan. These results support the contention that glucokinase regulates the metabolic flux rate through the glycolytic chain in the pancreatic B-cell and thereby generates the signal for glucose-induced insulin secretion.

Glucokinase is concentrated in insulin-secretory granules of pancreatic B-cells

1999 ◽  
Vol 112 (1) ◽  
pp. 35-40 ◽  
Author(s):  
Y. Toyoda ◽  
S. Yoshie ◽  
H. Shironoguchi ◽  
I. Miwa
Keyword(s):  
B Cells ◽  
Secretory Granules ◽  
Pancreatic B Cells

Fast insulin secretion reflects exocytosis of docked granules in mouse pancreatic B-cells

2002 ◽  
Vol 444 (1-2) ◽  
pp. 43-51 ◽  
Author(s):  
Charlotta S. Olofsson ◽  
Sven O. Göpel ◽  
Sebastian Barg ◽  
Juris Galvanovskis ◽  
Xiaosong Ma ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
B Cells ◽  
Pancreatic B Cells ◽  
Fast Insulin

Chronic Lymphocytic Leukemia B Cells Express Functional CXCR4 Chemokine Receptors That Mediate Spontaneous Migration Beneath Bone Marrow Stromal Cells

Blood ◽  
1999 ◽  
Vol 94 (11) ◽  
pp. 3658-3667 ◽  
Author(s):  
Jan A. Burger ◽  
Meike Burger ◽  
Thomas J. Kipps
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
Pertussis Toxin ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Actin Polymerization ◽  
Kinase Inhibitor ◽  
Critical Role ◽  
Lymphocytic Leukemia ◽  
Marrow Stromal Cells

Abstract Chemokines play a central role for lymphocyte trafficking and homing. The mechanisms that direct the tissue localization of B cells from patients with chronic lymphocytic leukemia (B-CLL) are unknown. We found that CLL B cells express functional CXCR4 receptors for the chemokine stromal cell-derived factor-1 (SDF-1), as demonstrated by receptor endocytosis, calcium mobilization, and actin polymerization assays. Moreover, CLL B cells displayed chemotaxis to this chemokine that could be inhibited by monoclonal antibodies (MoAbs) against CXCR4, pertussis toxin, or Wortmannin, a phosphatidylinositol 3-kinase inhibitor. That this chemotaxis may be involved in the homing of CLL cells is argued by studies in which CLL B cells were cocultured with a murine marrow stromal cell line that secretes SDF-1. Within 2 hours, CLL B cells spontaneously migrated beneath such stromal cells in vitro (pseudoemperipolesis). This migration could be inhibited by pretreatment of CLL B cells with anti-CXCR4 MoAbs, SDF-1, or pertussis-toxin. Furthermore, we noted strong downmodulation of CXCR4 on CLL B cells that migrated into the stromal cell layer. These findings demonstrate that the chemokine receptor CXCR4 on CLL B cells plays a critical role for heterotypic adherence to marrow stromal cells and provide a new mechanism to account for the marrow infiltration by neoplastic B cells.

scholarly journals Regulated Exocytosis in Neuroendocrine Cells: A Role for Subplasmalemmal Cdc42/N-WASP-induced Actin Filaments

2004 ◽  
Vol 15 (2) ◽  
pp. 520-531 ◽  
Author(s):  
Stéphane Gasman ◽  
Sylvette Chasserot-Golaz ◽  
Magali Malacombe ◽  
Michael Way ◽  
Marie-France Bader
Keyword(s):  
Plasma Membrane ◽  
Membrane Trafficking ◽  
Actin Polymerization ◽  
Cell Activation ◽  
Secretory Granules ◽  
Small Gtpases ◽  
Neuroendocrine Cells ◽  
Actin Dynamics ◽  
Cell Periphery ◽  
Regulated Exocytosis

In neuroendocrine cells, actin reorganization is a prerequisite for regulated exocytosis. Small GTPases, Rho proteins, represent potential candidates coupling actin dynamics to membrane trafficking events. We previously reported that Cdc42 plays an active role in regulated exocytosis in chromaffin cells. The aim of the present work was to dissect the molecular effector pathway integrating Cdc42 to the actin architecture required for the secretory reaction in neuroendocrine cells. Using PC12 cells as a secretory model, we show that Cdc42 is activated at the plasma membrane during exocytosis. Expression of the constitutively active Cdc42L61 mutant increases the secretory response, recruits neural Wiskott-Aldrich syndrome protein (N-WASP), and enhances actin polymerization in the subplasmalemmal region. Moreover, expression of N-WASP stimulates secretion by a mechanism dependent on its ability to induce actin polymerization at the cell periphery. Finally, we observed that actin-related protein-2/3 (Arp2/3) is associated with secretory granules and that it accompanies granules to the docking sites at the plasma membrane upon cell activation. Our results demonstrate for the first time that secretagogue-evoked stimulation induces the sequential ordering of Cdc42, N-WASP, and Arp2/3 at the interface between granules and the plasma membrane, thereby providing an actin structure that makes the exocytotic machinery more efficient.

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