scholarly journals Ca2+ entry into PC12 cells initiated by ryanodine receptors or inositol 1,4,5-trisphosphate receptors

1998 ◽  
Vol 329 (2) ◽  
pp. 349-357 ◽  
Author(s):  
L. Deborah BENNETT ◽  
D. Martin BOOTMAN ◽  
J. Michael BERRIDGE ◽  
R. Timothy CHEEK
Keyword(s):  
Endoplasmic Reticulum ◽  
Pc12 Cells ◽  
Ryanodine Receptors ◽  
Half Time ◽  
Free Medium ◽  
Excitable Cells ◽  
Inositol 1,4,5 Trisphosphate ◽  
Insp3 Receptors ◽  
Universal Mechanism ◽  
Stimulation Of

Capacitative Ca2+ entry (CCE) is a universal mechanism for refilling intracellular Ca2+ stores in electrically non-excitable cells. The situation in excitable cells is less clear, however, since they may rely on other entry mechanisms for Ca2+-store refilling. In the present study we investigated CCE in intact PC12 cells, using acetylcholine to bring about activation of InsP3 receptors (InsP3Rs), caffeine to activate ryanodine receptors (RyRs) and thapsigargin to inhibit sarco/endoplasmic reticulum Ca2+-ATPase pumps. We found that depletion of the InsP3-, caffeine- or thapsigargin-sensitive stores promoted Ca2+ entry, suggesting that stimulation of either InsP3Rs or RyRs can activate CCE. The CCE pathways activated by InsP3Rs, RyRs and thapsigargin appeared to be independent at least in part, since their effects were found to be additive. However, CCE triggered by caffeine, acetylcholine or thapsigargin progressively diminished with time. The decay of CCE caused by one agent also inhibited subsequent responses to the others, suggesting that some component of the CCE pathway is common to all intracellular Ca2+ stores. The magnitude of CCE stimulated by InsP3Rs or RyRs was related to the size of the stores; the InsP3-sensitive store was smaller than the RyR-sensitive store and triggered a smaller entry component. However, both stores filled with a similar half time (about 1 min), and both could be filled more rapidly by depolarization-induced Ca2+ entry through voltage-operated channels. A significant basal Ca2+ influx was apparent in PC12 cells. The basal entry component may be under the control of the InsP3-sensitive Ca2+ store, since short incubations in Ca2+-free medium depleted this store.

scholarly journals Effect of inositol 1,4,5-trisphosphate receptor stimulation on mitochondrial [Ca2+] and secretion in chromaffin cells

2002 ◽  
Vol 365 (2) ◽  
pp. 451-459 ◽  
Author(s):  
Mayte MONTERO ◽  
Maria Teresa ALONSO ◽  
Almudena ALBILLOS ◽  
Inmaculada CUCHILLO-IBÁÑEZ ◽  
Román OLIVARES ◽  
...  
Keyword(s):  
Chromaffin Cells ◽  
Ryanodine Receptors ◽  
Catecholamine Secretion ◽  
Mitochondrial Inhibitors ◽  
High K ◽  
Inositol 1,4,5 Trisphosphate ◽  
Insp3 Receptors ◽  
Voltage Dependent ◽  
Trisphosphate Receptor ◽  
Stimulation Of

Ca2+ uptake by mitochondria is a potentially important buffering system able to control cytosolic [Ca2+]. In chromaffin cells, we have shown previously that stimulation of either Ca2+ entry or Ca2+ release via ryanodine receptors triggers large increases in mitochondrial [Ca2+] ([Ca2+]M) approaching the millimolar range, whose blockade dramatically enhances catecholamine secretion [Montero, Alonso, Carnicero, Cuchillo-Ibañez, Albillos, Garcia, Carcia-Sancho and Alvarez (2000) Nat. Cell Biol. 2, 57–61]. In the present study, we have studied the effect of stimulation of inositol 1,4,5-trisphosphate (InsP3) receptors using histamine. We find that histamine produces a heterogeneous increase in [Ca2+]M, reaching peak levels at approx. 1μM in 70% of the mitochondrial space to several hundred micromolar in 2–3% of mitochondria. Intermediate levels were found in the rest of the mitochondrial space. Single-cell imaging experiments with aequorin showed that the heterogeneity had both an intercellular and a subcellular origin. Those mitochondria responding to histamine with increases in [Ca2+]M much greater than 1μM (30%) were the same as those that also responded with large increases in [Ca2+]M following stimulation with either high-K+ medium or caffeine. Blocking mitochondrial Ca2+ uptake with protonophores or mitochondrial inhibitors also enhanced catecholamine secretion induced by histamine. These results suggest that some InsP3 receptors tightly co-localize with ryanodine receptors and voltage-dependent Ca2+ channels in defined subplasmalemmal functional units designed to control secretion induced by different stimuli.

scholarly journals Caffeine-induced inhibition of inositol(1,4,5)-trisphosphate-gated calcium channels from cerebellum.

1994 ◽  
Vol 5 (1) ◽  
pp. 97-103 ◽  
Author(s):  
I Bezprozvanny ◽  
S Bezprozvannaya ◽  
B E Ehrlich
Keyword(s):  
Lipid Bilayers ◽  
Inhibitory Action ◽  
Single Channel ◽  
Ryanodine Receptors ◽  
Single Channels ◽  
Ca Channels ◽  
Open Time ◽  
Inositol 1,4,5 Trisphosphate ◽  
Insp3 Receptors ◽  
Intracellular Ca

Effects of the xanthine drug caffeine on inositol (1,4,5)-trisphosphate (InsP3)-gated calcium (Ca) channels from canine cerebellum were studied using single channels incorporated into planar lipid bilayers. Caffeine, used widely as an agonist of ryanodine receptors, inhibited the activity of InsP3-gated Ca channels in a noncooperative fashion with half-inhibition at 1.64 mM caffeine. The frequency of channel openings was decreased more than threefold after addition of 5 mM caffeine; there was only a small effect on mean open time of the channels, and the single channel conductance was unchanged. Increased InsP3 concentration overcame the inhibitory action of caffeine, but caffeine did not reduce specific [3H]InsP3 binding to the receptor. The inhibitory action of caffeine on InsP3 receptors suggests that the action of caffeine on the intracellular Ca pool must be interpreted with caution when both ryanodine receptors and InsP3 receptors are present in the cell.

scholarly journals Modeling the role of endoplasmic reticulum-mitochondria microdomains in calcium dynamics

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Arash Moshkforoush ◽  
Baarbod Ashenagar ◽  
Nikolaos M. Tsoukias ◽  
B. Rita Alevriadou
Keyword(s):  
Endoplasmic Reticulum ◽  
Vascular Endothelial Cells ◽  
Cell Model ◽  
Vascular Endothelial ◽  
Calcium Dynamics ◽  
Mitochondrial Membranes ◽  
Excitable Cells ◽  
Closed Cell ◽  
Stimulation Of

AbstractUpon inositol trisphosphate (IP3) stimulation of non-excitable cells, including vascular endothelial cells, calcium (Ca2+) shuttling between the endoplasmic reticulum (ER) and mitochondria, facilitated by complexes called Mitochondria-Associated ER Membranes (MAMs), is known to play an important role in the occurrence of cytosolic Ca2+ concentration ([Ca2+]Cyt) oscillations. A mathematical compartmental closed-cell model of Ca2+ dynamics was developed that accounts for ER-mitochondria Ca2+ microdomains as the µd compartment (besides the cytosol, ER and mitochondria), Ca2+ influx to/efflux from each compartment and Ca2+ buffering. Varying the distribution of functional receptors in MAMs vs. the rest of ER/mitochondrial membranes, a parameter called the channel connectivity coefficient (to the µd), allowed for generation of [Ca2+]Cytoscillations driven by distinct mechanisms at various levels of IP3 stimulation. Oscillations could be initiated by the transient opening of IP3 receptors facing either the cytosol or the µd, and subsequent refilling of the respective compartment by Ca2+ efflux from the ER and/or the mitochondria. Only under conditions where the µd became the oscillation-driving compartment, silencing the Mitochondrial Ca2+ Uniporter led to oscillation inhibition. Thus, the model predicts that alternative mechanisms can yield [Ca2+]Cyt oscillations in non-excitable cells, and, under certain conditions, the ER-mitochondria µd can play a regulatory role.

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 Inositol 1,4,5-trisphosphate mobilizes intracellular Ca2+ from permeabilized insulin-secreting cells

1984 ◽  
Vol 223 (2) ◽  
pp. 467-473 ◽  
Author(s):  
T J Biden ◽  
M Prentki ◽  
R F Irvine ◽  
M J Berridge ◽  
C B Wollheim
Keyword(s):  
Endoplasmic Reticulum ◽  
Steady State ◽  
Plasma Membranes ◽  
Cell Protein ◽  
Mitochondrial Activity ◽  
Rinm5f Cells ◽  
Cation Composition ◽  
Inositol 1,4,5 Trisphosphate ◽  
Insulin Secreting Cells ◽  
Stimulation Of

A possible role in secretory processes is proposed for inositol 1,4,5-triphosphate (IP3), based upon investigations of the Ca2+ steady state maintained by ‘leaky’, insulin-secreting RINm5F cells. These cells had been treated with digitonin to permeabilize their plasma membranes and thereby ensure that only intracellular Ca2+ buffering mechanisms were active. When placed in a medium with a cation composition resembling that of the cytosol, cells rapidly took up Ca2+ as measured by a Ca2+-specific minielectrode. Two Ca2+ steady states were observed. A lower level of around 120nM required ATP-dependent Ca2+ uptake and was probably determined by the endoplasmic reticulum. The higher steady state (approx. 800 nM), seen only in the absence of ATP, was shown to be due to mitochondrial activity. IP3 specifically released Ca2+ accumulated in the ATP-dependent pool, but not from mitochondria, since Ca2+ release was demonstrated in the presence of the respiratory poison antimycin. The IP3-induced Ca2+ release was rapid, with 50% of the response being seen within 15s. The apparent Km was 0.5 microM and maximal concentrations of IP3 (2.5 microM) produced a peak Ca2+ release of 10 nmol/mg of cell protein, which was followed by re-uptake. A full Ca2+ response was seen if sequential pulses of 2.5 microM-IP3 were added at 20 min intervals, although there was a slight (less than 20%) attenuation if the intervening period was decreased to 10 min. These observations could be related to the rate of IP3 degradation which, in this system, corresponded to a 25% loss of added 32P label within 2 min, and a 75% loss within 20 min. The results suggest that IP3 might act as a link between metabolic, cationic and secretory events during the stimulation of insulin release.

Calcium signalling in early divergence of Metazoa: mechanisms involved in the control of muscle-like cell contraction in Hydra plagiodesmica

2019 ◽  
Vol 97 (9) ◽  
pp. 812-824 ◽  
Author(s):  
María Eugenia Alzugaray ◽  
María Victoria Gavazzi ◽  
Jorge Rafael Ronderos
Keyword(s):  
Endoplasmic Reticulum ◽  
Homo Sapiens ◽  
Ryanodine Receptors ◽  
Calcium Signalling ◽  
Cell Contraction ◽  
Inositol 1,4,5 Trisphosphate ◽  
Trisphosphate Receptor

Our laboratory has previously examined the effect of neuropeptides on the activity of the hypostome of the hydra Hydra plagiodesmica Dioni, 1968 (Cnidaria: Hydrozoa). These results showed that the hypostome, a structure extruded during feeding, responds to myoregulatory peptides and that this mechanism might be regulated by changes in the cytosolic levels of calcium (Ca2+). We analyse now the ways in which Ca2+ modulates hypostome activity during feeding. The use of calcium chelators confirms that Ca2+ is relevant in inducing hypostome extrusion. The assay of compounds that modulate the activity of Ca2+ channels in the endoplasmic reticulum suggests that, beyond the extracellular influx of calcium, intracellular sources of the ion are involved and might include both ryanodine receptors (RyR) and the inositol 1,4,5-trisphosphate receptor (IP3R). Bioinformatic searches based on sequences of RyR and IP3R of humans (Homo sapiens Linnaeus, 1758) show that IP3Rs are present in all groups analysed, including Fungi and Choanoflagellata. Although H. plagiodesmica responds to caffeine and ryanodine, which are known to modulate RyRs, this family of receptors seems not to be predicted in Cnidaria, suggesting that this phylum either lacks these kinds of channels or that they possess a different structure compared with those possessed by other Metazoa.

A novel Ca2+-induced Ca2+ release mechanism mediated by neither inositol trisphosphate nor ryanodine receptors

2002 ◽  
Vol 361 (3) ◽  
pp. 605-611 ◽  
Author(s):  
Frank WISSING ◽  
Edmund P. NEROU ◽  
Colin W. TAYLOR
Keyword(s):  
Ryanodine Receptors ◽  
Ruthenium Red ◽  
Release Mechanism ◽  
Temperature Sensitive ◽  
Half Time ◽  
Ip3 Receptors ◽  
Inositol 1,4,5 Trisphosphate ◽  
Before And After ◽  
Partial Inactivation ◽  
Ca2 Channels

Members of both major families of intracellular Ca2+ channels, ryanodine and inositol 1,4,5-trisphosphate (IP3) receptors, are stimulated by substantial increases in cytosolic free Ca2+ concentration ([Ca2+]c). They thereby mediate Ca2+-induced Ca2+ release (CICR), which allows amplification and regenerative propagation of intracellular Ca2+ signals. In permeabilized hepatocytes, increasing [Ca2+]c to 10μM stimulated release of 30±1% of the intracellular stores within 60s; the EC50 occurred with a free [Ca2+] of 170±29nM. This CICR was abolished at 2°C. The same fraction of the stores was released by CICR before and after depletion of the IP3-sensitive stores, and CICR was not blocked by antagonists of IP3 receptors. Ryanodine, Ruthenium Red and tetracaine affected neither the Ca2+ content of the stores nor the CICR response. Sr2+ and Ba2+ (EC50 = 166nM and 28μM respectively) mimicked the effects of increased [Ca2+] on the intracellular stores, but Ni2+ blocked the passive leak of Ca2+ without blocking CICR. In rapid superfusion experiments, maximal concentrations of IP3 or Ca2+ stimulated Ca2+ release within 80ms. The response to IP3 was complete within 2s, but CICR continued for tens of seconds despite a slow [half-time (t1/2) = 3.54±0.07s] partial inactivation. CICR reversed rapidly (t1/2 = 529±17ms) and completely when the [Ca2+] was reduced. We conclude that hepatocytes express a novel temperature-sensitive, ATP-independent CICR mechanism that is reversibly activated by modest increases in [Ca2+], and does not require IP3 or ryanodine receptors or reversal of the sarcoplasmic/endoplasmic-reticulum Ca2+-ATPase. This mechanism may both regulate the Ca2+ content of the intracellular stores of unstimulated cells and allow even small intracellular Ca2+ signals to be amplified by CICR.

scholarly journals Luminal Ca2+ regulates passive Ca2+ efflux from the intracellular stores of hepatocytes

1998 ◽  
Vol 334 (2) ◽  
pp. 431-435 ◽  
Author(s):  
Mike D. BEECROFT ◽  
Colin W. TAYLOR
Keyword(s):  
Endoplasmic Reticulum ◽  
Steady State ◽  
Specific Activity ◽  
Critical Threshold ◽  
Similar Amount ◽  
Half Time ◽  
Atp Level ◽  
Simultaneous Addition ◽  
Inositol 1,4,5 Trisphosphate ◽  
The Many

Ca2+ uptake into the intracellular stores of permeabilized hepatocytes was entirely dependent on ATP and substantially inhibited by either ionomycin or thapsigargin, although both were required for complete inhibition. Unidirectional efflux of 45Ca2+ after removal of ATP from cells loaded to steady state (1.60±0.12 nmol/106 cells) was monoexponential and occurred with a half-time of 103±10 s. However, the 45Ca2+ content of the stores did not return to their pre-ATP level, but reached a plateau at 0.12±0.04 nmol/106 cells. A similar amount of Ca2+ was trapped within the stores when Ca2+ uptake was prevented by thapsigargin and chelation of Ca2+; at all temperatures between 2 °C and 37 °C; and after stores had first been loaded with unlabelled Ca2+. Simultaneous addition of inositol 1,4,5-trisphosphate (InsP3) and inhibition of Ca2+ uptake reduced the amount of trapped Ca2+ to a level consistent with InsP3 rapidly and more completely emptying a fraction of the stores that could be only partially emptied by the passive leak. After dilution of the specific activity of the 45Ca2+ under conditions that maintained the steady-state activities of the pumps and leaks, the stores rapidly lost their entire 45Ca2+ content. We conclude that the channel responsible for mediating the leak of Ca2+ abruptly closes when the luminal [Ca2+] of the intracellular stores falls below a critical threshold corresponding to about 7% of their steady-state loading. Whereas InsP3 is capable of completely emptying a fraction of the stores, regulation of the passive leak by luminal [Ca2+] is likely to prevent it from completely emptying them; such regulation may ensure that the many other functions of Ca2+ within the endoplasmic reticulum are not compromised.

scholarly journals Muscarinic receptor-induced phosphoinositide hydrolysis at resting cytosolic Ca2+ concentration in PC12 cells.

1985 ◽  
Vol 100 (4) ◽  
pp. 1330-1333 ◽  
Author(s):  
L M Vicentini ◽  
A Ambrosini ◽  
F Di Virgilio ◽  
T Pozzan ◽  
J Meldolesi
Keyword(s):  
Pc12 Cells ◽  
Muscarinic Receptor ◽  
Receptor Activation ◽  
Phosphoinositide Hydrolysis ◽  
Free Medium ◽  
Inositol 1,4,5 Trisphosphate ◽  
The Relationship ◽  
Hydrolysis Of ◽  
Cells Cultured

In PC12 cells, cultured in the presence of nerve growth factor to increase their complement of muscarinic receptors, treatment with carbachol induces muscarinic receptor-dependent rises in free cytosolic Ca2+ as well as hydrolysis of membrane phosphoinositides. Experiments were carried out to clarify the relationship between these two receptor-triggered events. In particular, since inositol-1,4,5-trisphosphate (the hydrophilic metabolite produced by the hydrolysis of phosphatidylinositol-4,5-bisphosphate) is believed to mediate intracellularly the release of Ca2+ from nonmitochondrial store(s), it was important to establish whether it can be generated at resting cytoplasmic concentration of Ca2+ (approximately 0.1 microM). Cells incubated in Ca2+-free medium were depleted of their cytoplasmic Ca2+ stores by pretreatment with ionomycin. When these cells were then treated with carbachol, their cytosolic concentration of Ca2+ remained at the resting level, whereas inositol-1,4,5-trisphosphate generation was still markedly stimulated. Our results demonstrate that an increase in the concentration of cytosolic Ca2+ is not a necessary intermediate between receptor activation and phosphoinositide hydrolysis, and therefore support the second-messenger role of inositol-1,4,5-trisphosphate.

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