scholarly journals Characterization of the inositol 1,4,5-trisphosphate-induced calcium release from permeabilized endocrine cells and its inhibition by decavanadate and p-hydroxymercuribenzoate

1989 ◽  
Vol 262 (1) ◽  
pp. 83-89 ◽  
Author(s):  
K J Föhr ◽  
J Scott ◽  
G Ahnert-Hilger ◽  
M Gratzl
Keyword(s):  
Endocrine Cells ◽  
Calcium Release ◽  
Cell Types ◽  
Inhibitory Effect ◽  
Maximal Effect ◽  
Thiol Compounds ◽  
Inositol 1,4,5 Trisphosphate ◽  
Dependent Inhibition ◽  
Pheochromocytoma Pc12 ◽  
First Time

The inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ compartment of endocrine cells was studied with alpha-toxin- and digitonin-permeabilized rat insulinoma (RINA2) and rat pheochromocytoma (PC12) cells. The Ca2+ uptake was ATP-dependent, and submicromolar concentrations of IP3 specifically released the stored Ca2+. Half-maximal Ca2+ release was observed with 0.25-0.5 mumol of IP3/l, and the amount of Ca2+ released due to IP3 could be enhanced by additional loading of the Ca2+ compartment. Consecutive additions of the same concentration of IP3 for 1-2 h always released the same amount of Ca2+ without desensitization, providing an ideal basis to further characterize the IP3-induced Ca2+ release. Here we describe for the first time a reversible inhibitory effect of decavanadate on the IP3-induced Ca2+ release. Among the vanadium species tested (decavanadate, oligovanadate and monovanadate), only decavanadate was inhibitory, with a half-maximal effect at 5 mumol/l in both cell types. The effect of decavanadate could be overcome by increasing the amount of sequestered Ca2+ or added IP3. Decavanadate did not affect the ATP-driven Ca2+ uptake but oligovanadate was inhibitory on Ca2+ uptake. p-Hydroxymercuribenzoate (pHMB) at concentrations between 10 and 30 mumol/l also inhibited the Ca2+ release due to IP3. Thiol compounds such as dithiothreitol (DTT; 1 mmol/l) added before pHMB removed all its inhibitory effect on the IP3-induced Ca2+ release, whereas the inhibition caused by decavanadate was unaffected by DTT. Thus, the decavanadate-dependent inhibition functions by a distinctly different mechanism than pHMB and could serve as a specific tool to analyse various aspects of the IP3-induced Ca2+ release within endocrine cells.

scholarly journals Autocrine down-regulation of glucocorticoid receptors by interleukin-11 in human osteoblast-like cell lines

2003 ◽  
Vol 177 (1) ◽  
pp. 109-117 ◽  
Author(s):  
A Dovio ◽  
ML Sartori ◽  
RG Masera ◽  
B Ceoloni ◽  
G Reimondo ◽  
...  
Keyword(s):  
Cell Lines ◽  
Glucocorticoid Receptors ◽  
Radioligand Binding ◽  
Inhibitory Effect ◽  
Human Osteoblast ◽  
Opposite Pattern ◽  
Dependent Inhibition ◽  
High Concentrations ◽  
First Time ◽  
Interleukin 11

It has recently been suggested that interleukin (IL)-11 plays a role in the pathogenesis of glucocorticoid (GC)-induced osteoporosis. IL-11 belongs to the gp130 cytokine family, which includes also IL-6. We have previously investigated GC-IL-6 interplay, showing that GC inhibits IL-6 release and IL-6 up-regulates GC receptor (GR) numbers in the human osteoblast-like cell lines Saos-2 and MG-63, which constitutively have an opposite pattern of expression for GR, IL-11, IL-6, alkaline phosphatase and osteoprotegerin (OPG). The aim of this study was to investigate GC-IL-11 interplay in the same two cell lines. First, cells were incubated with cortisol (0.01-1 microM) for 20 h in the presence and in the absence of a known IL-11 secretagogue (IL-1beta); cell media were assayed for IL-11 by ELISA. Secondly, cells were incubated with IL-11 (0.1-100 ng/ml) or specific anti-IL-11 monoclonal antibody for 20 h, and then assayed for GR by a radioligand binding assay. Similar to IL-6, both constitutive and IL-1beta-inducible IL-11 release were dose-dependently inhibited by cortisol (P<0.01); at variance with IL-6, exogenous IL-11 dose-dependently decreased GR numbers in MG-63 cells (P<0.05), while anti-IL-11 antibody significantly increased GR numbers in both cell lines (P<0.05). IL-11-induced reduction of GR in MG-63 cells was confirmed by Western blot analysis. While exerting opposite effects on GR numbers, neither IL-6 nor IL-11 significantly modified GC-dependent inhibition of OPG release. Our data indicate that even physiological concentrations of cortisol negatively modulate IL-11 secretion and demonstrate, for the first time, an inhibitory effect of the cytokine on GR. Thus, the concept of autocrine-paracrine loops that modulate GC action and involve gp130 cytokines is corroborated. These loops could have clinical relevance for the dynamics of bone loss in patients given GC and having high concentrations of these cytokines in the bone microenvironment.

scholarly journals Control of inositol 1,4,5-trisphosphate-induced Ca2+ release by cytosolic Ca2+

1995 ◽  
Vol 306 (2) ◽  
pp. 445-451 ◽  
Author(s):  
M D Bootman ◽  
L Missiaen ◽  
J B Parys ◽  
H De Smedt ◽  
R Casteels
Keyword(s):  
Binding Site ◽  
Cell Types ◽  
Inhibitory Effects ◽  
Activation Curve ◽  
Hormonal Stimulation ◽  
A7r5 Cells ◽  
Inositol 1,4,5 Trisphosphate ◽  
Insp3 Receptors ◽  
Dependent Inhibition ◽  
Stimulation Of

The synergistic action of cytosolic Ca2+ and inositol 1,4,5-trisphosphate (InsP3) in releasing intracellular Ca2+ stores has been suggested to be responsible for the complex intracellular Ca2 signals observed during hormonal stimulation of many cell types. However, the ability of cytosolic Ca2+ to potentiate Ca2+ release has recently been questioned because of the observed inhibitory effects of Ca2+ chelators used in previous studies. In the present study, EGTA and BAPTA [1,2-bis-(2-amino-phenoxy)ethane- NNN'N′-tetra-acetic acid] poorly inhibited InsP3-induced Ca2+ release from permeabilized A7r5 smooth-muscle cells. Additionally, stimulatory effects of cytosolic and luminal Ca2+ were observed either in the complete absence of Ca2+ chelator or at constant Ca(2+)-free chelator concentration. These data suggest that potentiation of InsP3-induced Ca2+ release by Ca2+ in A7r5 cells reflects an interaction between Ca2+ and InsP3 receptors, rather than a decrease in chelator-dependent inhibition. The EC50 for activation of InsP3-induced Ca2+ release by cytosolic Ca2+ was unaffected by ATP, or by changing InsP3 concentration, although InsP3-induced Ca2+ release became less sensitive to the inhibitory effects of cytosolic Ca2+ as the InsP3 concentration was elevated. Increasing H+ or Mg2+ concentration shifted the Ca(2+)-activation curve towards higher Ca2+ concentrations. These data suggest that, in addition to the InsP3-binding site, the affinity of the Ca(2+)-binding site(s) on InsP3 receptors can be modulated by intracellular cations.

scholarly journals Novel Regulation of Calcium Inhibition of the Inositol 1,4,5-trisphosphate Receptor Calcium-release Channel

2003 ◽  
Vol 122 (5) ◽  
pp. 569-581 ◽  
Author(s):  
Don-On Daniel Mak ◽  
Sean M.J. McBride ◽  
Nataliya B. Petrenko ◽  
J. Kevin Foskett
Keyword(s):  
Patch Clamp ◽  
Calcium Release ◽  
Critical Role ◽  
Cell Types ◽  
Dominant Negative ◽  
Bathing Solution ◽  
Calcium Release Channel ◽  
Release Channel ◽  
Calmodulin Antagonist ◽  
Inositol 1,4,5 Trisphosphate

The inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R), a Ca2+-release channel localized to the endoplasmic reticulum, plays a critical role in generating complex cytoplasmic Ca2+ signals in many cell types. Three InsP3R isoforms are expressed in different subcellular locations, at variable relative levels with heteromultimer formation in different cell types. A proposed reason for this diversity of InsP3R expression is that the isoforms are differentially inhibited by high cytoplasmic free Ca2+ concentrations ([Ca2+]i), possibly due to their different interactions with calmodulin. Here, we have investigated the possible roles of calmodulin and bath [Ca2+] in mediating high [Ca2+]i inhibition of InsP3R gating by studying single endogenous type 1 InsP3R channels through patch clamp electrophysiology of the outer membrane of isolated Xenopus oocyte nuclei. Neither high concentrations of a calmodulin antagonist nor overexpression of a dominant-negative Ca2+-insensitive mutant calmodulin affected inhibition of gating by high [Ca2+]i. However, a novel, calmodulin-independent regulation of [Ca2+]i inhibition of gating was revealed: whereas channels recorded from nuclei kept in the regular bathing solution with [Ca2+] ∼400 nM were inhibited by 290 μM [Ca2+]i, exposure of the isolated nuclei to a bath solution with ultra-low [Ca2+] (&lt;5 nM, for ∼300 s) before the patch-clamp experiments reversibly relieved Ca2+ inhibition, with channel activities observed in [Ca2+]i up to 1.5 mM. Although InsP3 activates gating by relieving high [Ca2+]i inhibition, it was nevertheless still required to activate channels that lacked high [Ca2+]i inhibition. Our observations suggest that high [Ca2+]i inhibition of InsP3R channel gating is not regulated by calmodulin, whereas it can be disrupted by environmental conditions experienced by the channel, raising the possibility that presence or absence of high [Ca2+]i inhibition may not be an immutable property of different InsP3R isoforms. Furthermore, these observations support an allosteric model in which Ca2+ inhibition of the InsP3R is mediated by two Ca2+ binding sites, only one of which is sensitive to InsP3.

Synthetic atrial peptide inhibits intracellular calcium release in smooth muscle

1986 ◽  
Vol 250 (1) ◽  
pp. C171-C174 ◽  
Author(s):  
K. D. Meisheri ◽  
C. J. Taylor ◽  
H. Saneii
Keyword(s):  
Smooth Muscle ◽  
Calcium Release ◽  
Rabbit Aorta ◽  
Inhibitory Effect ◽  
Physiological Salt Solution ◽  
Phasic Contraction ◽  
Dependent Inhibition ◽  
Isolated Rabbit Aorta ◽  
Dose Dependent Inhibition ◽  
45Ca Efflux

The effects of a synthetic atrial peptide (atriopeptin II; AP II) on the agonist-induced intracellular Ca2+ release was examined in the isolated rabbit aorta. The agonist-induced phasic contraction in a Ca2+-free physiological salt solution containing 2 mM ethyleneglycol-bis(beta-aminoethyl-ether)-N,N'-tetraacetic acid (EGTA-PSS) was used as an indicator of the intracellular Ca2+ release. The addition of AP II (10(-9)-10(-7) M) for 15 min to the tissue during the EGTA-PSS exposure caused a dose-dependent inhibition of norepinephrine (NE; 10(-6) M)-induced phasic contraction. The half-maximal inhibiting concentration of AP II was 3 X 10(-9) M, with 10(-7) M AP II causing 91% inhibition. This was confirmed by studying the inhibitory effect of AP II (10(-7) M) on NE-stimulated 45Ca efflux. Furthermore, the internal Ca2+ release by histamine (10(-5) M) and caffeine (25 mM), both of which share this internal Ca2+ pool with NE, was also inhibited by AP II. Thus AP II appears to be a potent inhibitor of the intracellular Ca2+ release that is utilized by various agonists for the activation of vascular smooth muscle. This may be an important mechanism by which AP II produces relaxation of blood vessels.

scholarly journals The effect of external calcium and pH on inositol trisphosphate-mediated calcium release from cerebellum microsomal fractions

1989 ◽  
Vol 258 (1) ◽  
pp. 261-265 ◽  
Author(s):  
S K Joseph ◽  
H L Rice ◽  
J R Williamson
Keyword(s):  
Calcium Release ◽  
Feedback Inhibition ◽  
Inhibitory Effect ◽  
Scatchard Analysis ◽  
Alkaline Ph ◽  
Apparent Affinity ◽  
Rat Cerebellum ◽  
Low Concentrations ◽  
Inositol 1,4,5 Trisphosphate ◽  
External Calcium

Binding of D-myo-inositol 1,4,5-trisphosphate (InsP3) to rat cerebellum membranes has previously been shown to be stimulated by alkaline pH and inhibited by low concentrations of Ca2+ [Worley, Baraban, Suppatopone, Wilson & Snyder (1987) J. Biol. Chem. 262, 12132-12136]. In the present study, Scatchard analysis of InsP3 binding to cerebellum microsomes indicates that the effects of Ca2+ and pH are exerted through changes in the apparent affinity of the receptor without effects on maximal binding. The influence of extravesicular Ca2+ and pH on InsP3-mediated 45Ca2+ release was investigated. Extravesicular Ca2+ inhibited InsP3-mediated Ca2+ release. The inhibitory effect of Ca2+ was most marked when a sub-optimal concentration of InsP3 was used. An increase in extravesicular pH produced a decrease in the concentration of InsP3 that yielded half-maximal Ca2+ release. Regulation of the affinity of the InsP3 receptor by Ca2+ and pH can qualitatively account for the observed effects of these factors on InsP3-mediated Ca2+ release. Feedback inhibition of InsP3 binding by Ca2+ could provide a mechanism to generate Ca2+ oscillations, particularly under hormonal conditions that produce sub-optimal elevations of InsP3 concentration.

Modulatory effect of glucose on VIP-induced gastric somatostatin release

1988 ◽  
Vol 254 (6) ◽  
pp. E756-E759 ◽  
Author(s):  
R. Schmid ◽  
V. Schusdziarra ◽  
M. Classen
Keyword(s):  
Cell Function ◽  
Inhibitory Effect ◽  
Maximal Effect ◽  
Inhibitory Effects ◽  
Glucose Levels ◽  
Normal Glucose ◽  
D Cell ◽  
First Time ◽  
Effect Of Glucose ◽  
Fasted Rats

The present study was designed to examine the effect of increasing perfusate glucose concentrations on vasoactive intestinal peptide (VIP)-induced somatostatin (SLI) release from the isolated rat stomach. The stomach of overnight-fasted rats was perfused with Krebs-Ringer buffer containing 100, 150, or 200 mg/dl glucose, respectively. VIP was administered at 10(-12), 10(-11), 10(-9), and 10(-8) M. At a normal glucose concentration of 100 mg/dl, VIP at doses of 10(-12), 10(-11), and 10(-9) M elicited a small inhibitory effect on SLI release by 200-300 pg/min (P less than 0.01). As reported previously at 10(-8) M, VIP stimulated gastric SLI secretion by 500 pg/min (P less than 0.01). Increasing perfusate glucose to 150 mg/dl resulted in a stimulation of SLI release by all four concentrations of VIP with a maximal effect at 10(-9) M. During 200 mg/dl glucose, VIP had no effect in concentrations below 10(-9) M, and only the two highest doses (10(-9) and 10(-8) M) stimulated SLI release significantly. In the absence of VIP, glucose had no effect on gastric SLI release. In conclusion, the present data demonstrate for the first time that at normal glucose levels VIP has not only stimulatory but also inhibitory effects on gastric SLI, and second, a modest elevation of glucose has a modulatory effect on gastric D-cell function.

scholarly journals NAADP induces pH changes in the lumen of acidic Ca2+ stores

2007 ◽  
Vol 402 (2) ◽  
pp. 301-310 ◽  
Author(s):  
Anthony J. Morgan ◽  
Antony Galione
Keyword(s):  
Nicotinic Acid ◽  
Direct Effect ◽  
Sea Urchin ◽  
Cell Types ◽  
Receptor Activation ◽  
Fluorescence Measurements ◽  
Low Concentrations ◽  
Inositol 1,4,5 Trisphosphate ◽  
Intracellular Organelles ◽  
First Time

NAADP (nicotinic acid–adenine dinucleotide phosphate)-induced Ca2+ release has been proposed to occur selectively from acidic stores in several cell types, including sea urchin eggs. Using fluorescence measurements, we have investigated whether NAADP-induced Ca2+ release alters the pHL (luminal pH) within these acidic stores in egg homogenates and observed their prompt, concentration-dependent alkalinization by NAADP (but not β-NAD+ or NADP). Like Ca2+ release, the pHL change was desensitized by low concentrations of NAADP suggesting it was secondary to NAADP receptor activation. Moreover, this was a direct effect of NAADP upon the acidic stores and not secondary to increases in cytosolic Ca2+ as it was not mimicked by IP3 (inositol 1,4,5-trisphosphate), cADPR (cyclic adenine diphosphoribose), ionomycin, thapsigargin or by direct addition of Ca2+, and was not blocked by EGTA. The results of the present study further support acidic stores as targets for NAADP and for the first time reveal an adjunct role for NAADP in regulating the pHL of intracellular organelles.

Ryanodine-induced calcium release from hepatic microsomes and permeabilized hepatocytes

1995 ◽  
Vol 268 (6) ◽  
pp. G1017-G1024 ◽  
Author(s):  
L. B. Lilly ◽  
J. L. Gollan
Keyword(s):  
Polyethylene Glycol ◽  
Functional Significance ◽  
Calcium Release ◽  
Cell Types ◽  
Endogenous Ligand ◽  
Cell Systems ◽  
Permeabilized Cells ◽  
Hepatic Microsomes ◽  
Inositol 1,4,5 Trisphosphate ◽  
Site Of Action

Inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] is a second messenger that releases Ca2+ from hepatocyte microsomes. The toxic alkaloid ryanodine modulates Ca2+ release via a receptor (RyR) identified in a variety of cell systems, but its regulation and functional significance in liver are undefined. Similarly, the role in hepatocyte Ca2+ regulation of adenosine 5'-cyclic diphosphate-ribose (cADPR), which is the putative endogenous ligand for RyR in other cell systems, has not been defined. Utilizing microsomes and permeabilized cells, we have investigated Ca2+ regulation in hepatocytes and, in particular, effects of ryanodine, cADPR, and other putative modulators on Ca2+ release and compared these with Ins(1,4,5)P3-induced Ca2+ release. Ryanodine at > or = 50 microM released 20% of microsomal Ca2+, and, in contrast to Ins(1,4,5)P3, no potentiation was observed with guanosine 5'-triphosphate and polyethylene glycol. Ins(1,4,5)P3-induced Ca2+ release was demonstrable after maximal ryanodine-induced Ca2+ release, suggesting that distinct Ca2+ stores are involved. cADPR (5 microM) did not induce Ca2+ release, alone or in combination with calmodulin or hepatic cytosol, nor did it influence ryanodine-induced release, in microsomes or permeabilized hepatocytes (in which ryanodine released 25% of the sequestered Ca2+). Ryanodine-induced Ca2+ release in microsomes was not influenced by 20 mM caffeine, which itself did not mobilize Ca2+, but was prevented by 500 microM tetracaine, which was shown to induce Ca2+ release. We conclude that ryanodine is capable of mobilizing Ca2+ in the hepatocyte from microsomal stores that are distinct from those that can be regulated by Ins(1,4,5)P3 but that cADPR has no such effect. These data suggest that cADPR does not serve as the endogenous ligand for RyR in liver cells or that the site of action of ryanodine in hepatocyte microsomes is distinct from that in other cell types.

scholarly journals Effects of adenine nucleotides on inositol 1,4,5-trisphosphate-induced calcium release in vascular smooth muscle cells.

1991 ◽  
Vol 98 (4) ◽  
pp. 681-698 ◽  
Author(s):  
M Iino
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Calcium Release ◽  
Adenine Nucleotides ◽  
Muscle Cells ◽  
Release Mechanism ◽  
Maximal Effect ◽  
Competitive Antagonism ◽  
Inositol 1,4,5 Trisphosphate ◽  
Intracellular Ca

Effects of adenine nucleotides on the inositol 1,4,5-trisphosphate (IP3)-induced Ca release (IICR) mechanism were studied in smooth muscle cells of the guinea pig portal vein. A microfluorometry method of fura-2 was used to measure Ca release from saponin-skinned thin muscle strips (width approximately 200 microns, thickness 50-70 microns, length 2-3 mm). About 80% of ionomycin-releasable Ca store was sensitive to IP3, of which approximately 20% was also sensitive to caffeine. The rate of Ca release by 0.1 microM IP3 depended biphasically on ATP concentration in the absence of Mg2+; it was dose-dependently enhanced by ATP up to approximately 0.5 mM, and above this concentration the enhancement became smaller. However, the decline of enhancement of the IICR at the higher ATP concentrations was absent at IP3 concentrations greater than 1 microM. This suggests competitive antagonism between IP3 and ATP. Clear effects of ATP were observed not only at pCa 7 or 8, where the Ca-induced Ca release was not activated, but after a ryanodine treatment to excise the functional compartment that possessed the Ca-induced Ca release mechanism. ATP had no effect on the rate of Ca leakage in the absence of IP3 even at pCa 5.5 after the ryanodine treatment. Therefore, ATP has direct biphasic effects on the IP3-induced Ca release mechanism. The Ca release induced by 0.1 microM IP3 at pCa 7 was potentiated not only by ATP, but by 0.5 mM ADP, AMP, or beta, gamma-methyleneadenosine 5'-triphosphate. 0.5 mM GTP had only a little effect on the IP3-induced Ca release. These results extend the functional similarities between Ca- and IP3-induced Ca release mechanisms in that adenine nucleotides enhance Ca release. Millimolar concentration of ATP, which is present physiologically, will shift the dose-response relation of IP3 toward the higher IP3 concentration and enhance the maximal effect of IP3. Thus, ATP is expected to assist the Ca release by higher concentrations of IP3 while having less effect on the Ca release by low levels of IP3. These effects of ATP may be important in the switching of Ca release from the intracellular Ca store by IP3.

scholarly journals Calcium release in HSY cells conforms to a steady-state mechanism involving regulation of the inositol 1,4,5-trisphosphate receptor Ca2+ channel by luminal [Ca2+].

1996 ◽  
Vol 132 (4) ◽  
pp. 607-616 ◽  
Author(s):  
A Tanimura ◽  
R J Turner
Keyword(s):  
Steady State ◽  
Calcium Release ◽  
Cell Types ◽  
Significant Shift ◽  
Ip3 Receptor ◽  
Permeabilized Cells ◽  
Fura 2 ◽  
State Process ◽  
Inositol 1,4,5 Trisphosphate ◽  
Trisphosphate Receptor

In many cell types, low concentrations of inositol 1,4,5-trisphosphate (IP3) release only a portion of the intracellular IP3-sensitive Ca2+ store, a phenomenon known as "quantal" Ca2+ release. It has been suggested that this effect is a result of reduced activity of the IP3-dependent Ca2+ channel with decreasing calcium concentration within the IP3-sensitive store ([Ca2+]s). To test this hypothesis, the properties of IP3-dependent Ca2+ release in single saponin-permeabilized HSY cells were studied by monitoring [Ca2+]s using the Ca(2+)-sensitive fluorescent dye mag-fura-2. In permeabilized cells, blockade of the sarco/ER Ca(2+)-ATPase pump in stores partially depleted by IP3 induced further Ca2+ release via an IP3-dependent route, indicating that Ca2+ entry via the sarco/ER Ca(2+)-ATPase pump had been balanced by Ca2+ loss via the IP3-sensitive channel before pump inhibition. IP3-dependent Mn2+ entry, monitored via quenching of luminal mag-fura-2 fluorescence, was readily apparent in filled stores but undetectable in Ca(2+)-depleted stores, indicating markedly reduced IP3-sensitive channel activity in the latter. Also consistent with reduced responsiveness of Ca(2+)-depleted stores to IP3, the initial rate of refilling of these stores was unaffected by the presence of 0.3 microM IP3, a concentration that was clearly effective in eliciting Ca2+ release from filled stores. Analysis of the rate of Ca2+ release at various IP3 concentrations indicated a significant shift of the IP3 dose response toward higher [IP3] with decreasing [Ca2+]s. We conclude that IP3-dependent Ca2+ release in HSY cells is a steady-state process wherein Ca2+ efflux via the IP3 receptor Ca2+ channel is regulated by [Ca2+]s, apparently via changes in the sensitivity of the channel to IP3.

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