scholarly journals Inositol trisphosphate analogues selective for types I and II inositol trisphosphate receptors exert differential effects on vasopressin-stimulated Ca2+ inflow and Ca2+ release from intracellular stores in rat hepatocytes

2004 ◽  
Vol 381 (2) ◽  
pp. 519-526 ◽  
Author(s):  
Roland B. GREGORY ◽  
Rachael HUGHES ◽  
Andrew M. RILEY ◽  
Barry V. L. POTTER ◽  
Robert A. WILCOX ◽  
...  
Keyword(s):  
Single Cells ◽  
Inositol Trisphosphate ◽  
Rat Hepatocytes ◽  
Small Region ◽  
Cell Periphery ◽  
Type I ◽  
Response Curves ◽  
Maximal Stimulation ◽  
Adenophostin A ◽  
Stimulation Of

Previous studies have shown that adenophostin A is a potent initiator of the activation of SOCs (store-operated Ca2+ channels) in rat hepatocytes, and have suggested that, of the two subtypes of Ins(1,4,5)P3 receptor predominantly present in rat hepatocytes [Ins(1,4,5)P3R1 (type I receptor) and Ins(1,4,5)P3R2 (type II receptor)], Ins(1,4,5)P3R1s are required for SOC activation. We compared the abilities of Ins(1,4,6)P3 [with higher apparent affinity for Ins(1,4,5)P3R1] and Ins(1,3,6)P3 and Ins(1,2,4,5)P4 [with higher apparent affinities for Ins(1,4,5)P3R2] to activate SOCs. The Ins(1,4,5)P3 analogues were microinjected into single cells together with fura 2, and dose–response curves for the activation of Ca2+ inflow and Ca2+ release from intracellular stores obtained for each analogue. The concentration of Ins(1,4,6)P3 which gave half-maximal stimulation of Ca2+ inflow was substantially lower than that which gave half-maximal stimulation of Ca2+ release. By contrast, for Ins(1,3,6)P3 and Ins(1,2,4,5)P3, the concentration which gave half-maximal stimulation of Ca2+ inflow was substantially higher than that which gave half-maximal stimulation of Ca2+ release. The distribution of Ins(1,4,5)P3R1 and Ins(1,4,5)P3R2 in rat hepatocytes cultured under the same conditions as those employed for the measurement of Ca2+ inflow and release was determined by immunofluorescence. Ins(1,4,5)-P3R1s were found predominantly at the cell periphery, whereas Ins(1,4,5)P3R2s were found at the cell periphery, the cell interior and nucleus. It is concluded that the idea that a small region of the endoplasmic reticulum enriched in Ins(1,4,5)P3R1 is required for the activation of SOCs is consistent with the present results for hepatocytes.

scholarly journals Evidence for the involvement of a small subregion of the endoplasmic reticulum in the inositol trisphosphate receptor-induced activation of Ca2+ inflow in rat hepatocytes

1999 ◽  
Vol 341 (2) ◽  
pp. 401-408 ◽  
Author(s):  
Roland B. GREGORY ◽  
Robert A. WILCOX ◽  
Leise A. BERVEN ◽  
Nicole C. R. VAN STRATEN ◽  
Gijs A. VAN DER MAREL ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Actin Cytoskeleton ◽  
Cytochalasin B ◽  
Rat Hepatocytes ◽  
Small Region ◽  
Cortical Actin ◽  
Isolated Rat Hepatocytes ◽  
Maximal Stimulation ◽  
Adenophostin A ◽  
Stimulation Of

The roles of a subregion of the endoplasmic reticulum (ER) and the cortical actin cytoskeleton in the mechanisms by which Ins(1,4,5)P3 induces the activation of store-operated Ca2+ channels (SOCs) in isolated rat hepatocytes were investigated. Adenophostin A, a potent agonist at Ins(1,4,5)P3 receptors, induced ER Ca2+ release and the activation of Ca2+ inflow. The concentration of adenophostin A that gave half-maximal stimulation of Ca2+ inflow (10 nM) was substantially lower than that (20 nM) which gave half-maximal ER Ca2+ release. A low concentration of adenophostin A (approx. 13 nM) caused near-maximal stimulation of Ca2+ inflow but only 20% of maximal ER Ca2+ release. Similar results were obtained using another Ins(1,4,5)P3-receptor agonist, 2-hydroxyethyl-α-D-glucopyranoside 2,3′,4′-trisphosphate. Anti-type-1 Ins(1,4,5)P3-receptor monoclonal antibody 18A10 inhibited vasopressin-stimulated Ca2+ inflow but had no observable effect on vasopressin-induced ER Ca2+ release. Treatment with cytochalasin B at a concentration that partially disrupted the cortical actin cytoskeleton inhibited Ca2+ inflow and ER Ca2+ release induced by vasopressin by 73 and 45%, respectively. However, it did not substantially affect Ca2+ inflow and ER Ca2+ release induced by thapsigargin or 13 nM adenophostin A, intracellular Ca2+ release induced by ionomycin or Ins(1,4,5)P3P4(5)-1-(2-nitrophenyl)ethyl ester [‘caged’ Ins(1,4,5)P3] or basal Ca2+ inflow. 1-(5-Chloronaphthalene-1-sulphonyl)homopiperazine, HCl (ML-9), an inhibitor of myosin light-chain kinase, also inhibited vasopressin-induced Ca2+ inflow and ER Ca2+ release by 53 and 44%, respectively, but had little effect on thapsigargin-induced Ca2+ inflow and ER Ca2+ release. Neither cytochalasin B nor ML-9 inhibited vasopressin-induced Ins(1,4,5)P3 formation. It is concluded that the activation of SOCs in rat hepatocytes induced by Ins(1,4,5)P3 requires the participation of a small region of the ER, which is distinguished from other regions of the ER by a different apparent affinity for Ins(1,4,5)P3 analogues and is associated with the plasma membrane through the actin skeleton. This conclusion is discussed briefly in relation to current hypotheses for the activation of SOCs.

scholarly journals Caffeine inhibits cytosolic calcium oscillations induced by noradrenaline and vasopressin in rat hepatocytes

1994 ◽  
Vol 301 (3) ◽  
pp. 737-744 ◽  
Author(s):  
L Combettes ◽  
B Berthon ◽  
M Claret
Keyword(s):  
Cyclic Amp ◽  
Single Cells ◽  
Rat Hepatocytes ◽  
Cytosolic Calcium ◽  
Inhibitory Effect ◽  
Calcium Oscillations ◽  
Direct Inhibition ◽  
Response Curves ◽  
Release Channel ◽  
Induced Changes

The effects of caffeine on agonist-induced changes in intracellular Ca2+ concentration ([Ca2+]i) were studied in single fura 2-loaded cells and suspensions of rat hepatocytes. In single cells, caffeine (5-10 mM) inhibited [Ca2+]i oscillations induced both by noradrenaline (0.1 microM) and by vasopressin (0.1 nM). Caffeine shifted the dose-response curves of the [Ca2+]i rise induced by vasopressin (0.5 to 2 nM) and noradrenaline (from 80 to 580 nM) in suspensions of liver cells loaded with quin2. This inhibitory effect of caffeine was not due to inhibition of phosphodiesterase enzymes and elevation of cyclic AMP levels, because application of 3-isobutyl-1-methylxanthine, forskolin or 8-bromo cyclic AMP had no inhibitory effect on the intracellular Ca2+ rise induced by inositol 1,4,5-trisphosphate (InsP3)-dependent agonists. We demonstrate that the inhibitory effect of caffeine may result from at least three actions of caffeine: (1) inhibition of receptor-stimulated InsP3 formation; (2) inhibition of agonist-stimulated Ca2+ influx; and (3) direct inhibition of the InsP3-sensitive Ca(2+)-release channel.

Cation-dependent vitamin D activation of human renal cortical guanylate cyclase

1984 ◽  
Vol 246 (1) ◽  
pp. E115-E120 ◽  
Author(s):  
D. L. Vesely ◽  
D. Juan
Keyword(s):  
Vitamin D ◽  
Guanylate Cyclase ◽  
Cellular Level ◽  
Cyclase Activity ◽  
Rat Tissues ◽  
Response Curves ◽  
Guanylate Cyclase Activity ◽  
Manganese Ion ◽  
Maximal Stimulation ◽  
Stimulation Of

The objective of this investigation was to determine whether physiological levels of vitamin D and its metabolites have part of their mechanisms of action through stimulation of guanylate cyclase (EC 4.6.1.2). These sterols enhanced both soluble and particulate guanylate cyclase activities as well as cGMP levels two- to threefold in human and rat tissues. At a concentration of 1 nM, 1,25(OH)2D3 greater than 25(OH)D3 greater than vitamin D3 greater than 24,25(OH)2D3 = 25,26(OH)2D3 = vitamin D2. Dose-response curves revealed that maximal stimulation of guanylate cyclase by these sterols was at 1 nM and that there was no augmented guanylate cyclase activity at 0.01 nM. The precursors of vitamin D, cholesterol and 7-dehydrocholesterol, had no effect on guanylate cyclase activity. The activation of guanylate cyclase activity by the vitamin D sterols required the presence of manganese ion. Calcium was not as efficient as manganese in optimizing basal or hormone-stimulated guanylate cyclase activity. Vitamin D and its metabolites failed to stimulate adenylate cyclase (EC 4.6.1.1) activity. The data in this investigation suggest that guanylate cyclase may play a role in the mechanism of action of vitamin D at the cellular level.

Angiotensin II stimulates guanylate cyclase activity in aorta, heart, and kidney

1981 ◽  
Vol 240 (4) ◽  
pp. E391-E393
Author(s):  
D. L. Vesely
Keyword(s):  
Angiotensin Ii ◽  
Mechanism Of Action ◽  
Guanylate Cyclase ◽  
Rat Aorta ◽  
Cellular Level ◽  
Cyclase Activity ◽  
Response Curves ◽  
Guanylate Cyclase Activity ◽  
Maximal Stimulation ◽  
Stimulation Of

The objective of the present investigation was to determine whether angiotensin II at physiological levels has part of its mechanism of action through stimulation of the activity of guanylate cyclase (EC 4.6.1.2), the enzyme that catalyzes the conversion of guanosine triphosphate to cyclic GMP. Angiotensin II enhanced guanylate cyclase activity three-to fivefold in rat aorta, heart, and kidney at a concentration of 1 nM. Dose-response curves revealed that near maximal stimulation of guanylate cyclase with angiotensin II was observed at a concentration as low as 10 pM. The guanylate cyclase cofactor manganese was necessary for the maximal enhancement of guanylate cyclase by angiotensin II. The data in this investigation suggest that guanylate cyclase may play a role in the mechanism of action of angiotensin II at the cellular level.

Flux through glycine cleavage system in isolated hepatocytes: effects of glucagon, cAMP, and calcium

1990 ◽  
Vol 68 (2) ◽  
pp. 543-546 ◽  
Author(s):  
Markandeya Jois ◽  
Beatrice Hall ◽  
Vaughn M. Collett ◽  
John T. Brosnan
Keyword(s):  
Acid Metabolism ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Liver Mitochondria ◽  
Glycine Cleavage System ◽  
Isolated Rat Hepatocytes ◽  
Maximal Stimulation ◽  
Glycine Cleavage ◽  
Camp Levels ◽  
Stimulation Of

The hepatic glycine cleavage system (GCS) is the principal route for the metabolism of glycine in mammals. Flux through the GCS in isolated rat hepatocytes was stimulated by about 100% by glucagon (10−7 M), forskolin (10−4 M), and dibutyryl cAMP (10−4 M). The stimulation of flux through the GCS by these agents was accompanied by marked elevation of cellular cAMP levels. A significant correlation was observed between increased cellular cAMP levels induced by glucagon and stimulation of flux through the GCS by glucagon. Exclusion of calcium from the incubation medium reduced the basal flux by 38%, but did not affect the degree of stimulation of flux through the GCS by glucagon. A single intraperitoneal injection of glucagon to rats prior to isolation of hepatocytes resulted in a 76% stimulation of flux through the GCS. These hepatocytes with stimulated flux through the GCS showed reduced sensitivity for further stimulation by glucagon. Half-maximal stimulation of flux through the GCS occurred at 3.8 ± 1.1 and 8.5 ± 1.4 nM glucagon in hepatocytes isolated from control and glucagon-injected rats, respectively. We conclude that cAMP is involved in the regulation of flux through the GCS by gluagon.Key words: amino acid, metabolism, liver, mitochondria, hormones.

scholarly journals Different receptors use inositol trisphosphate to mobilize Ca2+ from different intracellular pools

2000 ◽  
Vol 351 (3) ◽  
pp. 683-686 ◽  
Author(s):  
Alison D. SHORT ◽  
Gavin P. WINSTON ◽  
Colin W. TAYLOR
Keyword(s):  
Spatial Organization ◽  
Inositol Trisphosphate ◽  
Signalling Pathways ◽  
Free Medium ◽  
Hek 293 ◽  
Hek 293 Cells ◽  
Maximal Stimulation ◽  
293 Cells ◽  
Made In ◽  
Stimulation Of

In cells expressing different receptors linked to Ins(1,4,5)P3 formation, maximal stimulation of any one of them often releases all the Ins(1,4,5)P3-sensitive Ca2+ stores, suggesting that Ins(1,4,5)P3 is used similarly by many receptors. In single HEK-293 cells, ATP and carbamylcholine (CCh) stimulated Ca2+ release from intracellular stores via a pathway that was entirely dependent on Ins(1,4,5)P3. After stimulation with maximal concentrations of ATP or CCh in Ca2+-free medium, there was no response to a second stimulation with the same agonist, indicating that each agonist had emptied the Ins(1,4,5)P3-sensitive stores to which it had access. However, the Ca2+ release evoked by the second agonist was unaffected by prior stimulation with the first. We conclude that Ins(1,4,5)P3 mediates the effects of both receptors, but Ins(1,4,5)P3 is more versatile than hitherto supposed, because the spatial organization of the signalling pathways apparently allows Ins(1,4,5)P3 made in response to each agonist to interact with different Ins(1,4,5)P3 receptors.

GTP analogues stimulate inositol trisphosphate formation transiently in Dictyostelium

1987 ◽  
Vol 87 (4) ◽  
pp. 513-518
Author(s):  
G.N. Europe-Finner ◽  
P.C. Newell
Keyword(s):  
Cyclic Amp ◽  
G Proteins ◽  
Inositol Trisphosphate ◽  
Signal Transduction Pathway ◽  
Transduction Pathway ◽  
Permeabilized Cells ◽  
Maximal Stimulation ◽  
Gamma S ◽  
Stimulation Of

Permeabilization of amoebae of Dictyostelium discoideum with saponin was found not to uncouple the chemotactic cell surface cyclic AMP receptors from inositol trisphosphate (IP3) formation, and stimulation of permeabilized amoebae with 50 nM-cyclic AMP produced peaks of IP3 at 5, 15 and 30 s in a manner comparable to that seen previously in non-permeabilized cells. The possible involvement of a GTP-binding protein (G-protein) in this IP3 signal transduction pathway was investigated by studying the effects on such permeabilized amoebae of added GTP and non-hydrolysable GTP analogues. While GDP produced only very minor effects, stimulation of the amoebae (in the absence of added cyclic AMP) with GTP or the non-hydrolysable GTP analogues GTP gamma S (guanosine 5′-O-(3-thio-triphosphate] and Gpp(NH)p (5′-guanylylimidodiphosphate) induced transient formation of IP3 in an oscillatory manner, with peaks similar in magnitude and timing to those elicited by cyclic AMP. A dose-response curve for GTP gamma S indicated a concentration for half-maximal stimulation of approximately 8 microM. When tested at 300 s after addition of GTP gamma S, the basal level of IP3 was found to be twofold elevated with shallow (presumably asynchronous) oscillations still just discernible. The significance of the IP3 oscillations elicited by GTP and its analogues is discussed in relation to the mechanism of signal adaptation and the presumed role of G-proteins.

scholarly journals α-adrenergic stimulation of respiration in isolated rat hepatocytes

1983 ◽  
Vol 210 (3) ◽  
pp. 867-873 ◽  
Author(s):  
A Binet ◽  
M Claret
Keyword(s):  
Respiration Rate ◽  
Rat Hepatocytes ◽  
Adrenergic Stimulation ◽  
Isolated Rat Hepatocytes ◽  
Cell Responses ◽  
Maximal Stimulation ◽  
Lag Period ◽  
Dose Dependent ◽  
Isolated Rat ◽  
Stimulation Of

1. The alpha-adrenergic agonists noradrenaline (in the presence of beta-blocker) and phenylephrine cause a transient stimulation of the respiration in isolated rat hepatocytes. After a lag period of 12s, this activation first attains its maximal value (+24%) for about 1 min and then falls to a sustained value (+15%). The effect is blocked by the alpha-antagonists phenoxybenzamine and phentolamine. It is dose-dependent, with an half-maximal stimulation by 16 nM-noradrenaline, which is similar to that found for other cell responses to the hormone. 2. Vasopressin and ATP, which in common with alpha-agonists are believed to increase intracellular [Ca2+], induce similar activation in the respiration rate. 3. The alpha-adrenergic-mediated respiration depends on extracellular Ca2+. The activation is decreased or abolished when extracellular [Ca2+] is decreased by adding EGTA, or when the Ca2+ antagonists Mn2+ and La3+ are present in the incubation medium. 4. It is suggested that the activation of the mitochondrial respiration rate results from the increase in cytosolic Ca2+ concentration, presumably via Ca2+ influx or Ca2+ release from the plasma membrane or endoplasmic reticulum.

EFFECTS OF INCREASING DOSES OF PYROGLUTAMYL-HISTIDYL-PROLINE AMIDE ON SERUM THYROTROPHIN LEVELS IN NORMAL SUBJECTS

1972 ◽  
Vol 70 (1) ◽  
pp. 196-208 ◽  
Author(s):  
Bengt Karlberg ◽  
Sven Almqvist
Keyword(s):  
Normal Saline ◽  
Standard Dose ◽  
Normal Subjects ◽  
Response Curves ◽  
Clinical Observations ◽  
Trh Stimulation ◽  
Stimulation Tests ◽  
Serial Measurements ◽  
Serum Tsh ◽  
Stimulation Of

ABSTRACT The effects of the administration of normal saline in four normal subjects and the single iv injections of synthetic pyroglutamyl-histidyl-proline amide (TRH) in doses of 6.25, 12.5, 25, 50, 100, 200 and 400 μg in 12 healthy subjects were evaluated by clinical observations and serial measurements from −10 to + 360 minutes of serum TSH, PBI, STH, cholesterol, glucose and insulin. Normal saline and TRH 6.25 μg iv did not change the serum TSH level. The minimum iv dose of TRH increasing serum TSH within 10 minutes was 12.5 μg. Nine of 12 subjects gave maximal increases of serum TSH after TRH 100 μg and all after 200 and 400 μg. The time for the peak response varied with the dose from 15 to 60 minutes. The dose-response curves, average and individual, were complex and not linear. This was interpreted as a varying degree of stimulation of both pituitary synthesis and release of TSH by TRH. PBI changes were measured at 2 h and 6 h. Minimum dose for a significant increase of PBI was 12.5 μg and 6.25 μg of TRH for the respective times. No change in basal STH-levels occurred in 53 of 65 TRH-stimulation tests. Nine of the 12 changes in serum STH occurred in four subjects with varying basal STH-levels. No changes were found in serum cholesterol, glucose or insulin. Our results show that 50 μg of TRH can be used as a standard dose for the single iv stimulation of pituitary release of TSH. TRH stimulated both TSH and STH release in 18% of our tests.

Use of Flexible Materials with a Novel Pressure Driven Equibiaxial Cell Stretching Device for Mechanical Stimulation of Single Mammalian Cells

2003 ◽  
Vol 773 ◽  
Author(s):  
James D. Kubicek ◽  
Stephanie Brelsford ◽  
Philip R. LeDuc
Keyword(s):  
Cell Fate ◽  
Mechanical Stimulation ◽  
Mammalian Cells ◽  
Cellular Response ◽  
Single Cells ◽  
Complex Nature ◽  
Cellular Behavior ◽  
Cell Stretching ◽  
Stimulation Of ◽  
Pressure Driven

AbstractMechanical stimulation of single cells has been shown to affect cellular behavior from the molecular scale to ultimate cell fate including apoptosis and proliferation. In this, the ability to control the spatiotemporal application of force on cells through their extracellular matrix connections is critical to understand the cellular response of mechanotransduction. Here, we develop and utilize a novel pressure-driven equibiaxial cell stretching device (PECS) combined with an elastomeric material to control specifically the mechanical stimulation on single cells. Cells were cultured on silicone membranes coated with molecular matrices and then a uniform pressure was introduced to the opposite surface of the membrane to stretch single cells equibiaxially. This allowed us to apply mechanical deformation to investigate the complex nature of cell shape and structure. These results will enhance our knowledge of cellular and molecular function as well as provide insights into fields including biomechanics, tissue engineering, and drug discovery.

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