What Do You Know About IP6/IP-6/inositol Hexaphosphate/phytate/phytic Acid?

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ABSTRACT The GTPase Ras is a molecular switch engaged downstream of G-protein-coupled receptors and receptor tyrosine kinases that controls multiple cell-fate-determining signalling pathways. Ras signalling is frequently deregulated in cancer, underlying associated changes in cell phenotype. Although Ca2+ signalling pathways control some overlapping functions with Ras, and altered Ca2+ signalling pathways are emerging as important players in oncogenic transformation, how Ca2+ signalling is remodelled during transformation and whether it has a causal role remains unclear. We have investigated Ca2+ signalling in two human colorectal cancer cell lines and their isogenic derivatives in which the allele encoding oncogenic K-Ras (G13D) was deleted by homologous recombination. We show that agonist-induced Ca2+ release from the endoplasmic reticulum (ER) intracellular Ca2+ stores is enhanced by loss of K-RasG13D through an increase in the Ca2+ content of the ER store and a modification of the abundance of inositol 1,4,5-trisphosphate (IP3) receptor (IP3R) subtypes. Consistently, uptake of Ca2+ into mitochondria and sensitivity to apoptosis was enhanced as a result of K-RasG13D loss. These results suggest that suppression of Ca2+ signalling is a common response to naturally occurring levels of K-RasG13D, and that this contributes to a survival advantage during oncogenic transformation. KEY WORDS: K-R

RESEARCH ARTICLE Oncogenic K-Ras suppresses IP3-dependent Ca2+ release through remodelling of the isoform composition of IP3Rs and ER luminal Ca2+ levels in colorectal cancer cell lines Cristina Pierro, Simon J. Cook, Thomas C. F. Foets, Martin D. Bootman* and H. Llewelyn Roderick 2014

http://jcs.biologists.org/content/joces/127/7/1607.full.pdf
 
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Remodeling of Ca 2+ signaling in cancer: Regulation of inositol 1,4,5-trisphosphate receptors through oncogenes and tumor suppressors
Article (PDF Available)  in Advances in Biological Regulation 68 · December 2017 with 146 Reads
DOI: 10.1016/j.jbior.2017.12.001

Abstract

The calcium ion (Ca2+) is a ubiquitous intracellular signaling molecule that regulates diverse physiological and pathological processes, including cancer. Increasing evidence indicates that oncogenes and tumor suppressors regulate the Ca2+ transport systems. Inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) are IP3-activated Ca2+ release channels located on the endoplasmic reticulum (ER). They play pivotal roles in the regulation of cell death and survival by controlling Ca2+ transfer from the ER to mitochondria through mitochondria-associated ER membranes (MAMs). Optimal levels of Ca2+ mobilization to mitochondria are necessary for mitochondrial bioenergetics, whereas excessive Ca2+ flux into mitochondria causes loss of mitochondrial membrane integrity and apoptotic cell death. In addition to well-known functions on outer mitochondrial membranes, B-cell lymphoma 2 (Bcl-2) family proteins are localized on the ER and regulate IP3Rs to control Ca2+ transfer into mitochondria. Another regulatory protein of IP3R, IP3R-binding protein released with IP3 (IRBIT), cooperates with or counteracts the Bcl-2 family member depending on cellular states. Furthermore, several oncogenes and tumor suppressors, including Akt, K-Ras, phosphatase and tensin homolog (PTEN), promyelocytic leukemia protein (PML), BRCA1, and BRCA1 associated protein 1 (BAP1), are localized on the ER or at MAMs and negatively or positively regulate apoptotic cell death through interactions with IP3Rs and regulation of Ca2+ dynamics. The remodeling of Ca2+ signaling by oncogenes and tumor suppressors that interact with IP3Rs has fundamental roles in the pathology of cancers.

https://www.researchgate.net/public...ptors_through_oncogenes_and_tumor_suppressors
 
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Calcium Signaling VOL.1 NO.1 MARCH 2014 ISSN: 2373-1168 (Print) / 2373-1176 (Online) CS

Inositol 1, 4, 5-trisphosphate Receptor in Cancer: Good Cop or Bad Cop?

Aru Singh# , M.Sc., Megha Chagtoo# , M.Sc., and Bandana Chakravarti, Ph.D.*

Abstract:
Calcium signaling is very complex and intimately linked with cell survival and physiological form of cell death. Endoplasmic reticulum is the main intracellular storage of Ca2+ and release of Ca2+ is controlled by ER membrane localized IP3Rs and RYRs. These receptors regulate the transfer of Ca2+ from ER to mitochondria via transport protein on mitochondrial membrane which regulates cellular bioenergetics. Altered IP3R activity and/or the remodeling of IP3R-expression profiles may be exploited by cancer cells to promote growth and drug resistance. It can also affect mitochondrial bio-energetics and susceptibility to apoptotic stimuli, thereby enabling the survival of cells with oncogenic features. Three subtypes of IP3R (IP3R1, IP3R2, and IP3R3) share basic properties but differ in terms of regulation. To what extent they contribute to complex Ca2+ signaling in cancer cells remains largely unknown. Understanding the detailed molecular regulation of IP3Rs will be important to develop novel therapeutic strategies to target cancer cells through their deregulated Ca2+-signaling machinery. Here we will review the role of IP3R in different cancer as a biomarker or druggable target.

Source: https://pdfs.semanticscholar.org/6344/a865e593bf0dcfc4a2949987363eff3c60ce.pdf
 
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Cell Signalling: IP3 Receptors Channel Calcium into Cell Death
Current Biology, Volume 14, Issue 21, pR933–R935, 9 November 2004
Calcium Group, Laboratory of Molecular Signalling, Babraham Institute, Babraham, Cambridge CB2 4AT, UK
DOI: Redirecting
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C.Jane Hanson, Martin D. Bootman, Llewelyn Roderick
Email the author Martin D. Bootman

Abstract

There is substantial evidence that Ca2+ fluxes occur during most forms of apoptosis, and that inhibiting such fluxes protects cells from death. IP3 receptors – ligand-gated channels that release Ca2+ from intracellular stores – are emerging as key sites for regulation by pro- and anti-apoptotic factors.

The cytosolic Ca2+ concentration in resting cells is maintained at low levels (∼100 nM) by enzymes that translocate Ca2+ ions across the plasma membrane or into intracellular stores. Ca2+ uptake into mitochondria is another important way of removing cytosolic Ca2+ ions: by sequestering Ca2+ ions, mitochondria can modulate the kinetics and spatial dimensions of cellular Ca2+ signals [1]. Elevation of Ca2+ within the mitochondrial matrix activates citric acid cycle enzymes and thereby stimulates ATP production [2]. The cytosolic Ca2+ level is increased physiologically when cells are challenged with stimuli such as hormones, growth factors, depolarisation or mechanical deformation. The Ca2+ions can be released from intracellular stores, enter the cell across the plasma membrane, or a combination of the two. Cells employ a diverse range of messengers and channels to access these sources [3]. For mobilising Ca2+ stores, inositol 1,4,5-trisphosphate (IP3) receptors are a principal route in almost all cell types. The release of Ca2+ via IP3 receptors stimulates activities critical for life, such as post-fertilization Ca2+ oscillations [4], but under some conditions IP3 receptor-mediated Ca2+ signals are subverted to cause cell death.

IP3 receptors are large (∼1200 kDa) tetrameric proteins, each subunit of which projects an amino-terminal domain into the cytoplasm, their membrane-spanning carboxy-terminal regions forming an integral Ca2+ channel. IP3 binding by the amino-terminal domains causes a conformational change that promotes channel opening. Between the IP3 binding site and the transmembrane regions is a large stretch of amino acids where a significant proportion of regulatory interactions occur. Although IP3 is necessary to open native IP3 receptors, activation of these channels is complex and their open probability actually depends on the ambient Ca2+ concentration. Up to ∼500 nM, Ca2+ works synergistically with IP3 to activate IP3 receptors. At higher concentrations, cytosolic Ca2+ inhibits IP3 receptor opening. The inhibition of IP3receptors by Ca2+ is thought to be a crucial mechanism for terminating channel activity and thus preventing pathological Ca2+ rises.

IP3 Receptors in Apoptosis

A number of studies have shown that reducing IP3 receptor expression inhibits apoptosis. For example, DT40 cells (a chicken B lymphoma cell line) in which IP3 receptor expression was prevented were not only deficient in IP3-mediated Ca2+ signalling, but also substantially resistant to the apoptosis normally induced in response to B-cell receptor activation [5]. Similarly, Jurkat cells with reduced expression of IP3receptors showed only a modest activation of caspases 3 and 9 following engagement of the CD3 component of the T-cell receptor [6]. Caspases are aspartate-directed cysteine proteases responsible for a cascade of events that culminate in cell death.

Three IP3 receptor isoforms have been cloned and splice variants have been described, leading to the possibility that heteromultimeric channels are assembled with distinctive properties relating to their subunit content. At present, it is not clear whether different IP3 receptor isoforms have an equivalent role in apoptosis. In the studies using DT40 cells, there appeared to be redundancy between different IP3receptor isoforms. Cells where all three isoforms were knocked out showed the least death compared to cells missing either a single isoform or pairs of isoforms [5]. In contrast to this, some studies have shown that reduction of either type 1 or type 3 IP3 receptor abrogates apoptosis.

How Does Ca2+ Cause Apoptosis?

Transfer of Ca2+ between intracellular stores and mitochondria provides physiological control of respiration. But this Ca2+ cycle can also lead to cell death. If the matrix Ca2+ level rises too high, or if normal Ca2+ signals occur concurrently with production of molecules such as arachidonate or ceramide, then deleterious changes in mitochondrial structure may occur. In particular, mitochondria can swell and rupture or undergo permeability transition, thereby releasing several pro-apoptotic factors into the cytoplasm, such as cytochrome C, second mitochondrial activator of caspases (SMAC/Diablo) or apoptosis-inducing factor (AIF) [7]. This leads to the generation of the ‘apoptosome’ and activation of caspases from inactive zymogens. It is well established that Ca2+ released through IP3 receptors is sequestered by mitochondria [2]. Furthermore, it has been demonstrated that the flow of Ca2+ specifically from IP3 receptors can cause mitochondrial permeability transition and activate the apoptotic cascade [8].

In addition to directly feeding mitochondria with Ca2+, activation of IP3 receptors can stimulate a variety of Ca2+-sensitive enzymes that engage other apoptotic mechanisms. One clear target of IP3-mediated Ca2+ signals is the phosphatase calcineurin. This enzyme has many substrates, but in particular it can dephosphorylate the pro-apoptotic protein Bad causing it to translocate from the cytosol to mitochondria and thereby stimulate cytochrome C release. T cells deficient in IP3 receptors were found to be resistant to a variety of apoptotic stimuli; however, expression of a constitutively active form of calcineurin or pharmacological elevation of Ca2+ could by-pass the need for IP3-induced Ca2+ release and restore cell death [9].

How Are IP3 Receptors Activated during Apoptosis?

The physiological way of activating IP3 receptors, of course, is to provoke intracellular production of IP3. Natural IP3-generating agonists can induce apoptosis under some conditions. In the case of immature B cells, for example, apoptosis can be induced by IP3-mediated Ca2+signals arising from activation of the B-cell receptor. This is essential to establish immunological self-tolerance. In contrast, with mature B cells, B-cell receptor engagement provokes proliferation and antibody production. In a similar vein, application of an IP3-generating agonist to mouse embryonic fibroblasts enhanced apoptosis when it was applied concurrently with a pro-apoptotic stimulus such as arachidonate [10].

Recent studies have demonstrated that IP3 receptors can also be activated during apoptosis independently of the production of IP3. In the case of type 1 IP3 receptors, this may occur following proteolysis by activated caspase 3. The type 1 IP3 receptor isoform has a single highly conserved DEVD cleavage site, which is caspase 3 specific. Truncation of IP3 receptors by caspase 3 at this position removes a large portion of the protein, including the amino-terminal IP3-binding region and other regulatory domains. The part that remains in the endoplasmic reticulum (ER) membrane is a constitutively active channel that continuously leaks Ca2+[11].

The critical role of IP3 receptor cleavage by caspase 3 in apoptosis was recently demonstrated by Assefa et al.[12]. They used DT40 cells in which all three IP3 receptor isoforms had been knocked out so that the cells are insensitive to staurosporine: exogenous expression of normal type 1 IP3 receptors – but not mutant receptors that cannot be cut by caspase-3 – restored sensitivity to staurosporine-induced apoptosis [12]. Furthermore, expression of the ‘channel only’ domain produced following proteolysis of IP3 receptors in the triple knock-out DT40 cells also sensitised them to apoptosis.

Many pro-apoptotic stimuli, such as staurosporine, cause Ca2+ signals as part of their mechanism of killing. Buffering the Ca2+ rise can prevent, or at least delay, the onset of apoptosis [13]. The study by Assefa et al.[12] suggests that the Ca2+ elevation occurs because of IP3receptor cleavage. Consistent with this, the DT40 cells expressing normal type 1 IP3 receptors, but not the caspase-resistant mutant, displayed increased cytosolic Ca2+ following staurosporine addition. The Ca2+ rise was blocked by a specific caspase 3 inhibitor [12]. These data reveal that type 1 IP3 receptors are critical targets for caspase cleavage, and the subsequent unregulated release of Ca2+ accelerates cell death.

In an alternative model for IP3 receptor activation during apoptosis that is not reliant on caspase cleavage, cytochrome C released from mitochondria is suggested to control Ca2+ movement through IP3 receptors. Cytochrome C was shown to bind IP3 receptors and reduce the Ca2+-dependent inhibition of channel opening [14]. Within a cell undergoing apoptosis, this generates a positive feedback loop, whereby Ca2+ stimulates cytochrome C release and vice versa. The significance of this amplification mechanism was demonstrated by expressing a peptide corresponding to the carboxy-terminal portion of IP3 receptors, where cytochrome C binds: this peptide buffers cytochrome C as it emerges from mitochondria and prevents its interaction with functional IP3 receptors. As predicted, the carboxy-terminal IP3 receptor fragment acted in a dominant-negative fashion and substantially reduced both the amplitude of Ca2+ signals and release of cytochrome C following addition of staurosporine [14].

These studies all point to a crucial role of IP3 receptors in the generation of Ca2+ signals that drive cells into apoptosis. But it is unclear which model accounts for the activation of Ca2+ release. The schemes involving caspase 3 cleavage and cytochrome C binding appear to be mutually exclusive, in that IP3 receptors cleaved by caspase 3 are constitutively open and so presumably do not require cytochrome C binding. On the other hand, cytochrome C only regulates channels that can still bind IP3, but caspase 3 cleavage removes the IP3 binding site. It is possible that caspase 3 and cytochrome C have a synergystic action, work sequentially or target different IP3 populations.

IP3 receptors are by no means the only mechanism by which cellular Ca2+ can be elevated during cell death. Several other types of channel are known to transport Ca2+ and promote apoptosis. In particular, members of the TRPM family – melastatin-related transient receptor potential cation channels – underlie Ca2+ fluxes and cell death in neurons. And IP3 receptors are not the only targets for caspases. It has been demonstrated that the Ca2+ATPases on the plasma membrane are cleaved by caspases, thus preventing normal homeostasis and consequently causing Ca2+ elevation [15]. Damage to cellular membranes can cause a leak of Ca2+ into the cytoplasm, which often leads to the activation of Ca2+-dependent cysteine endopeptidases known as calpains. These proteases have been shown to activate caspases, cause cytochrome C release and promote cell death.

Interaction with the Anti-Apoptotic Protein Bcl-2
As Ca2+ signals emanating from the ER via IP3 receptors are critical for induction of cell death, regulating Ca2+ release is an obvious way for cells to control apoptotic signalling. Consistent with this, the anti-apoptotic protein Bcl-2 has been shown to reduce ER Ca2+ release, decrease mitochondrial Ca2+ uptake and abrogate cell death. Bcl-2 is a small integral membrane protein that localizes to both mitochondria and the ER [16] and so is present at the key locations where Ca2+ transport occurs. It has been proposed that Bcl-2 decreases the Ca2+content of the ER and thereby reduces the flux of Ca2+ from IP3 receptors to mitochondria. But whilst it is clear that manipulating ER luminal Ca2+ concentration can alter the sensitivity of cells to apoptosis [17], whether Bcl-2 works via this mechanism is controversial [18].

An alternative scheme by which Bcl-2 might regulate IP3 receptor activity was recently highlighted by the observation of a direct interaction between these proteins [19]. Expression of Bcl-2 in a T-cell line was found to inhibit Ca2+ signals in response to a membrane-permeant IP3analogue, without altering the ER Ca2+ content. Furthermore, the open probability of purified IP3 receptors incorporated into planar lipid bilayers was reduced by addition of recombinant Bcl-2. The direct coupling of these proteins within cells was substantiated using bi-directional co-immunoprecipitation of IP3 receptors (isoforms 1 and 3) and Bcl-2. Where Bcl-2 binds to IP3 receptors, and how it alters channel activity, are not known. Although the experiments in planar lipid bilayers suggest that the interaction is direct, it is plausible that within cells Bcl-2 also acts to scaffold other proteins to IP3 receptors. For example, it has been proposed that Bcl-2 docks calcineurin with IP3receptors [20]. By regulating the phosphorylation status of IP3 receptors, calcineurin could modulate Ca2+ release.

Without the participation of IP3 receptors, cells are either resistant to pro-apoptotic stimulation or cell death is dramatically slowed. The precise mechanism by which IP3 receptors are recruited during apoptosis is uncertain. It is clear, however, that Ca2+ accelerates the rate of apoptosis, and that IP3 receptors provide an amplification mechanism by which sustained Ca2+ signals can arise (Figure 1).

figure 1.jpg

Figure 1
Some of the interactions that lead to recruitment of IP3 receptors during apoptosis.

The positive feedback between IP3 receptor-mediated Ca2+ release and mitochondria underlies the generation of Ca2+ signals that accelerate the rate of cell death. The apoptosis-inducing cycle of Ca2+ between IP3 receptors and mitochondria can be initiated by a variety of mechanisms, including non-specific entry of Ca2+ following membrane damage.

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References

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© 2004 Elsevier Ltd. Published by Elsevier Inc.
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Source: https://www.cell.com/fulltext/S0960-9822(04)00810-3
 
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Oncotarget. 2016 Sep 20;7(38):61403-61418. doi: 10.18632/oncotarget.8968.
Sulforaphane-induced apoptosis involves the type 1 IP3 receptor.


Abstract

In this study we show that anti-tumor effect of sulforaphane (SFN) is partially realized through the type 1 inositol 1,4,5-trisphosphate receptor (IP3R1). This effect was verified in vitro on three different stable cell lines and also in vivo on the model of nude mice with developed tumors. Early response (6 hours) of A2780 ovarian carcinoma cells to SFN treatment involves generation of mitochondrial ROS and increased transcription of NRF2 and its downstream regulated genes including heme oxygenase 1, NAD(P)H:quinine oxidoreductase 1, and KLF9. Prolonged SFN treatment (24 hours) upregulated expression of NRF2 and IP3R1. SFN induces a time-dependent phosphorylation wave of HSP27. Use of IP3R inhibitor Xestospongin C (Xest) attenuates both SFN-induced apoptosis and the level of NRF2 protein expression. In addition, Xest partially attenuates anti-tumor effect of SFN in vivo. SFN-induced apoptosis is completely inhibited by silencing of IP3R1 gene but only partially blocked by silencing of NRF2; silencing of IP3R2 and IP3R3 had no effect on these cells. Xest inhibitor does not significantly modify SFN-induced increase in the rapid activity of ARE and AP1 responsive elements. We found that Xest effectively reverses the SFN-dependent increase of nuclear content and decrease of reticular calcium content. In addition, immunofluorescent staining with IP3R1 antibody revealed that SFN treatment induces translocation of IP3R1 to the nucleus. Our results clearly show that IP3R1 is involved in SFN-induced apoptosis through the depletion of reticular calcium and modulation of transcription factors through nuclear calcium up-regulation.

Source: Sulforaphane-induced apoptosis involves the type 1 IP3 receptor. - PubMed - NCBI
 
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Genet Mol Res. 2015 Jun 26;14(2):6929-42. doi: 10.4238/2015.June.26.1.
Mechanisms of cytotoxicity induced by the anesthetic isoflurane: the role of inositol 1,4,5-trisphosphate receptors.
Zhai WH1, Zhao J2, Huo SP3, Chen XG3, Li YD1, Zhang ZL1, Yu LL4, Song S5, Wang QJ6.
Author information
1 Department of Anesthesiology, Affiliated Hospital of Jining Medical University, Jining, China.
2 Department of Cell Biology, School of Basic Medical Sciences, Hebei Medical University, Shijiazhuang, Hebei, China.
3 Department of Anesthesiology, Third Hospital of Hebei Medical University, Shijiazhuang, Hebei, China.
4 Department of Anesthesiology, Cangzhou Central Hospital, Cangzhou, China.
5 Department of Anesthesiology, Yuhuanding Hospital, Yantai, China.
6 Department of Anesthesiology, Third Hospital of Hebei Medical University, Shijiazhuang, Hebei, China [email protected].
Abstract
Isoflurane can induce widespread cytotoxicity. We hypothesized that isoflurane induces apoptosis partly by causing excessive calcium release from the endoplasmic reticulum (ER) via direct activation of inositol 1,4,5-trisphosphate receptors (IP3R). Rat pheochromocytoma cells cultured for seven days with nerve growth factor were divided into four groups: control group (C), IP3R antagonist group (X), isoflurane group (I) and isoflurane + IP3R antagonist group (I+X). Groups I and I+X were treated with 1 MAC isoflurane for 12 h. Groups X and I+X were pretreated with IP3R antagonist. Annexin V/PI apoptosis and TUNEL assays were performed to evaluate cell apoptosis. TEM was used to observe changes in cell ultrastructure. Changes in calcium concentration ([Ca(2+)]i) in the cytoplasm were measured by flow cytometry. RT-PCR was performed to evaluate IP3R mRNA expression. TEM showed that isoflurane treatment altered cell ultrastructure. Compared to group C, cell apoptosis rate and [Ca(2+)]i increased in groups I and I+X (P < 0.05). Compared to group C, IP3R mRNA expression was lower in group X and higher in group I (P < 0.05). Compared to group X, cell apoptosis rate, [Ca(2+)]i and IP3R mRNA expression increased in groups I and I+X (P < 0.05). Compared to group I, cell apoptosis rate, [Ca(2+)]i and IP3R mRNA expression decreased in group I+X (P < 0.05). These results suggest that exposure to 1 MAC isoflurane for 12 h causes excessive calcium release partly by direct activation of IP3R on the ER membrane and triggers cell apoptosis.
 
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Role of subcellular calcium redistribution in regulating apoptosis and autophagy in cadmium-exposed primary rat proximal tubular cells
Fei Liu, Zi-Fa Li, Zhen-Yong Wang, Lin Wang

College of Animal Science and Veterinary Medicine, Shandong Agricultural University, Daizong Road No. 61, Tai'an 271018, People's Republic of China
Laboratory Animal Center of Shandong University of Traditional Chinese Medicine, Jinan 250355, People's Republic of China
Received 13 June 2016, Revised 30 August 2016, Accepted 13 September 2016, Available online 14 September 2016.

Abstract

Ca2 + signaling plays a vital role in regulating apoptosis and autophagy. We previously proved that cytosolic Ca2 + overload is involved in cadmium (Cd)-induced apoptosis in rat proximal tubular (rPT) cells, but the source of elevated cytosolic Ca2 + concentration ([Ca2 +]c) and the effect of potential subcellular Ca2 + redistribution on apoptosis and autophagy remain to be elucidated. Firstly, data showed that Cd-induced elevation of [Ca2 +]c was primarily generated intracellularly. Moreover, elevations of [Ca2 +]c and mitochondrial Ca2 + concentration ([Ca2 +]mit) with depletion of endoplasmic reticulum (ER) Ca2 + levels ([Ca2 +]ER) were revealed in Cd-treated rPT cells, but this subcellular Ca2 +redistribution was significantly suppressed by 2-Aminoethoxydiphenyl borate (2-APB). Elevated inositol 1,4,5-trisphosphate (IP3) levels with up-regulated IP3 receptor (IP3R) protein levels were shown in Cd-exposed cells, confirming that IP3R-mediated ER Ca2 +release results in the elevation of [Ca2 +]c. Up-regulated sequestosome 1 (p62) protein levels and autophagic flux assay demonstrated that Cd impaired autophagic degradation, while N-acetylcysteine (NAC) markedly attenuated Cd-induced p62 and microtubule-associated protein 1 light chain 3-II (LC3-II) accumulation, implying that the inhibition of autophagic flux was due to oxidative stress. Furthermore, pharmacological modulation of [Ca2 +]c with 1,2-Bis (2-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid acetoxymethyl ester (BAPTA-AM) and 2-APB alleviated Cd-mediated apoptosis, inhibition of autophagic degradation and subsequent cytotoxicity, while thapsigargin (TG) had the opposite regulatory effect on them. In summary, cytosolic calcium overload originated from IP3R-mediated ER Ca2 + release has a negative impact on Cd nephrotoxicity through its promotion of apoptosis and inhibition of autophagic flux.

Source: Role of subcellular calcium redistribution in regulating apoptosis and autophagy in cadmium-exposed primary rat proximal tubular cells - ScienceDirect
 
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Exp Ther Med. 2018 Apr;15(4):3791-3800. doi: 10.3892/etm.2018.5902. Epub 2018 Feb 28.
Chronic ethanol exposure induces SK-N-SH cell apoptosis by increasing N-methyl-D-aspartic acid receptor expression and intracellular calcium.
Wang H1, Wang X1, Li Y2, Yu H1, Wang C1, Feng C1, Xu G1, Chen J1, You J1, Wang P1, Wu X1, Zhao R1, Zhang G1.
Author information
1 Department of Forensic Pathology, School of Forensic Medicine, China Medical University, Shenyang, Liaoning 110122, P.R. China.
2 No.1 English Department, School of Fundamental Sciences, China Medical University, Shenyang, Liaoning 110122, P.R. China.
It has been identified that chronic ethanol exposure damages the nervous system, particularly neurons. There is scientific evidence suggesting that neuronal loss caused by chronic ethanol exposure has an association with neuron apoptosis and intracellular calcium oscillation is one of the primary inducers of apoptosis. Therefore, the present study aimed to investigate the inductive effects of intracellular calcium oscillation on apoptosis in SK-N-SH human neuroblastoma cells and the protective effects of the N-methyl-D-aspartic acid receptor (NMDAR) antagonist, memantine, on SK-N-SH cell apoptosis caused by chronic ethanol exposure. SK-N-SH cells were treated with 100 mM ethanol and memantine (4 µM) for 2 days. Protein expression of NR1 was downregulated by RNA interference (RNAi). Apoptosis was detected by Annexin V/propidium iodide (PI) double-staining and flow cytometry and cell viability was detected using an MTS kit. Fluorescence dual wavelength spectrophotometry was used to determine the intracellular calcium concentration and the levels of NR1 and caspase-3 were detected using western blotting. NR1 mRNA levels were also detected using qPCR. It was found that chronic ethanol exposure reduced neuronal cell viability and caused apoptosis of SK-N-SH cells, and the extent of damage in SK-N-SH cells was associated with ethanol exposure concentration and time. In addition, chronic ethanol exposure increased the concentration of intracellular calcium in SK-N-SH cells by inducing the expression of NMDAR, resulting in apoptosis, and memantine treatment reduced ethanol-induced cell apoptosis. The results of the present study indicate that the application of memantine may provide a novel strategy for the treatment of alcoholic dementia.

Source: Chronic ethanol exposure induces SK-N-SH cell apoptosis by increasing N-methyl-D-aspartic acid receptor expression and intracellular calcium. - PubMed - NCBI
 
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J Mol Neurosci. 2017 Aug;62(3-4):402-411. doi: 10.1007/s12031-017-0948-3. Epub 2017 Jul 20.
Memantine Can Reduce Ethanol-Induced Caspase-3 Activity and Apoptosis in H4 Cells by Decreasing Intracellular Calcium.
Wang X1, Chen J1, Wang H1, Yu H1, Wang C1, You J1, Wang P1, Feng C1, Xu G1, Wu X1, Zhao R1, Zhang G2.
Author information
1 Department of Forensic Pathology, School of Forensic Medicine, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, Liaoning Province, People's Republic of China.
2 Department of Forensic Pathology, School of Forensic Medicine, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, Liaoning Province, People's Republic of China. [email protected].
Abstract
Caspase-3 activation and apoptosis are associated with various neurodegenerative disorders. Calcium activation is an important factor in promoting apoptosis. We, therefore, assessed the role of intracellular calcium in ethanol-induced activation of caspase-3 in H4 human neuroglioma cells and the protective effect of the NMDA receptor antagonist, memantine, on ethanol-induced apoptosis in H4 cells. H4 cells were treated with 100 mM EtOH (in culture medium) for 2 days. For interaction studies, cells were treated with memantine (4 μM), EDTA (1 mM), or BAPTA-AM (10 μM) before treatment with EtOH. Knockdown of the gene encoding the NR1 subunit of the NMDA receptor was performed using RNAi. Apoptosis was detected by Annexin V-FITC/PI staining and flow cytometry. Cell viability was detected using an MTS cell proliferation kit. Fluorescence dual wavelength spectrophotometry was used to determine the intracellular calcium concentration. The levels of NR1, caspase-3, IP3R1, and SERCA1 proteins were detected by western blotting. NR1, IP3R1, and SERCA1 mRNA levels were detected by qPCR. We observed increased expression of NR1, IP3R1, SERCA1, and increased intracellular levels of calcium ions in H4 cells exposed to ethanol. In addition, the calcium chelators, EDTA and BAPTA, and RNAi disruption of the NMDA receptor reduced ethanol-induced caspase-3 activation in H4 cells. Memantine treatment reduced the ethanol-induced increase of intracellular calcium, caspase-3 activation, apoptosis, and the ethanol-induced decrease in cell viability. Our results indicate that ethanol-induced caspase-3 activation and apoptosis are likely to be dependent on cytosolic calcium levels and that they can be reduced by memantine treatment.

Source: Memantine Can Reduce Ethanol-Induced Caspase-3 Activity and Apoptosis in H4 Cells by Decreasing Intracellular Calcium. - PubMed - NCBI
 
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General anesthetic isoflurane modulates inositol 1,4,5-trisphosphate receptor calcium channel opening.
Joseph JD1, Peng Y, Mak DO, Cheung KH, Vais H, Foskett JK, Wei H.
Author information
1 From the Department of Anesthesiology and Critical Care (J.D.J., Y.P., H.W.) and Department of Physiology (D.-O.D.M., K.-H.C., H.V., J.K.F.), Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania. Current address: Department of Physiology, University of Hong Kong, Hong Kong, China (K.-H.C.).
Abstract
BACKGROUND:
Pharmacological evidence suggests that inhalational general anesthetics induce neurodegeneration in vitro and in vivo through overactivation of inositol trisphosphate receptor (InsP3R) Ca-release channels, but it is not clear whether these effects are due to direct modulation of channel activity by the anesthetics.

METHODS:
Using single-channel patch clamp electrophysiology, the authors examined the gating of rat recombinant type 3 InsP3R (InsP3R-3) Ca-release channels in isolated nuclei (N = 3 to 15) from chicken lymphocytes modulated by isoflurane at clinically relevant concentrations in the absence and presence of physiological levels of the agonist inositol 1,4,5-trisphosphate (InsP3). The authors also examined the effects of isoflurane on InsP3R-mediated Ca release from the endoplasmic reticulum and changes in intracellular Ca concentration ([Ca]i).

RESULTS:
Clinically relevant concentrations (approximately 1 minimal alveolar concentration) of the commonly used general anesthetic, isoflurane, activated InsP3R-3 channels with open probability similar to channels activated by 1 µM InsP3 (Po ≈ 0.2). This isoflurane modulation of InsP3R-3 Po depended biphasically on [Ca]i. Combination of isoflurane with subsaturating levels of InsP3 in patch pipettes resulted in at least two-fold augmentations of InsP3R-3 channel Po compared with InsP3 alone. These effects were not noted in the presence of saturating [InsP3]. Application of isoflurane to DT40 cells resulted in a 30% amplification of InsP3R-mediated [Ca]i oscillations, whereas InsP3-induced increase in [Ca]i and cleaved caspase-3 activity were enhanced by approximately 2.5-fold.

CONCLUSION:
These results suggest that the InsP3R may be a direct molecular target of isoflurane and plays a role in the mechanisms of anesthetic-mediated pharmacological or neurotoxic effects.

Source: General anesthetic isoflurane modulates inositol 1,4,5-trisphosphate receptor calcium channel opening. - PubMed - NCBI
 
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Pflugers Arch. 2014 Jul;466(7):1329-42. doi: 10.1007/s00424-013-1366-1. Epub 2013 Oct 10.
Sodium/calcium exchanger is upregulated by sulfide signaling, forms complex with the β1 and β3 but not β2 adrenergic receptors, and induces apoptosis.
Markova J1, Hudecova S, Soltysova A, Sirova M, Csaderova L, Lencesova L, Ondrias K, Krizanova O.
Author information
1 Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences (SAS), Vlarska 5, 833 34, Bratislava, Slovak Republic.
Abstract
Hydrogen sulfide (H2S) as a novel gasotransmitter regulates variety of processes, including calcium transport systems. Sodium calcium exchanger (NCX) is one of the key players in a regulation calcium homeostasis. Thus, the aims of our work were to determine effect of sulfide signaling on the NCX type 1 (NCX1) expression and function in HeLa cells, to investigate the relationship of β-adrenergic receptors with the NCX1 in the presence and/or absence of H2S, and to determine physiological importance of this potential communication. As a H2S donor, we used morpholin-4-ium-4-methoxyphenyl(morpholino) phosphinodithioate-GYY4137. We observed increased levels of the NCX1 mRNA, protein, and activity after 24 h of GYY4137 treatment. This increase was accompanied by elevated cAMP due to the GYY4137 treatment, which was completely abolished, when NCX1 was silenced. Increased cAMP levels would point to upregulation of β-adrenergic receptors. Indeed, GYY4137 increased expression of β1 and β3 (but not β2) adrenergic receptors. These receptors co-precipitated, co-localized with the NCX1, and induced apoptosis in the presence of H2S. Our results suggest that sulfide signaling plays a role in regulation of the NCX1, β1 and β3 adrenergic receptors, their co-localization, and stimulation of apoptosis, which might be of a potential importance in cancer treatment.

Source: Sodium/calcium exchanger is upregulated by sulfide signaling, forms complex with the β1 and β3 but not β2 adrenergic receptors, and induces apoptosis. - PubMed - NCBI
 
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Prog Neuropsychopharmacol Biol Psychiatry. 2013 Dec 2;47:156-61. doi: 10.1016/j.pnpbp.2013.05.009. Epub 2013 May 28.
Dual effects of neuroprotection and neurotoxicity by general anesthetics: role of intracellular calcium homeostasis.
Wei H1, Inan S.
Author information
1
Department of Anesthesiology and Critical Care, University of Pennsylvania, 305 John Morgan Building, 3620 Hamilton Walk, Philadelphia, PA 19104, USA. Electronic address: [email protected].
Abstract
Although general anesthetics have long been considered neuroprotective, there are growing concerns about neurotoxicity. Preclinical studies clearly demonstrated that commonly used general anesthetics are both neuroprotective and neurotoxic, with unclear mechanisms. Recent studies suggest that differential activation of inositol 1,4,5-trisphosphate receptors, a calcium release channel located on the membrane of endoplasmic reticulum (ER), play important role on determining the fate of neuroprotection or neurotoxicity by general anesthetics. General anesthetics at low concentrations for short duration are sublethal stress factors which induce endogenous neuroprotective mechanisms and provide neuroprotection via adequate activation of InsP3R and moderate calcium release from ER. On the other hand, general anesthetics at high concentrations for prolonged duration are lethal stress factors which induce neuronal damage by over activation of InsP3R and excessive and abnormal Ca(2+) release from ER. This review emphasizes the dual effects of both neuroprotection and neurotoxicity via differential regulation of intracellular Ca(2+) homeostasis by commonly used general anesthetics and recommends strategy to maximize neuroprotective but minimize neurotoxic effects of general anesthetics.

Source: Dual effects of neuroprotection and neurotoxicity by general anesthetics: role of intracellular calcium homeostasis. - PubMed - NCBI
 
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Phosphorylated K-Ras limits cell survival by blocking Bcl-xL sensitization of inositol trisphosphate receptors
Pamela J. Sung, Frederick D. Tsai, Horia Vais, Helen Court, Jun Yang, Nicole Fehrenbacher, J. Kevin Foskett, and Mark R. Philips
PNAS December 17, 2013. 110 (51) 20593-20598; Phosphorylated K-Ras limits cell survival by blocking Bcl-xL sensitization of inositol trisphosphate receptors

  1. Edited* by Joseph Schlessinger, Yale University School of Medicine, New Haven, CT, and approved November 6, 2013 (received for review April 15, 2013)
Significance
K-Ras is mutated more often than any other oncogene, making the protein and the pathways it regulates attractive targets for anticancer drug discovery. We have shown that phosphorylation of serine 181 in the membrane-targeting region of K-Ras causes the protein to translocate from plasma membrane to intracellular membranes. Translocation is associated with toxicity but the mechanism has remained undefined. Here we show that phospho–K-Ras associates with inositol trisphosphate receptors (IP3Rs) on the endoplasmic reticulum (ER) and thereby blocks one of the prosurvival activities of Bcl-xL, which is to sensitize IP3Rs and thereby allow constitutive transfer of calcium from ER to mitochondria where it is required for efficient respiration. This pathway could be exploited to limit the oncogenic activity of mutant K-Ras.

Abstract
K-Ras4B is targeted to the plasma membrane by a farnesyl modification that operates in conjunction with a polybasic domain. We characterized a farnesyl-electrostatic switch whereby protein kinase C phosphorylates K-Ras4B on serine 181 in the polybasic region and thereby induces translocation from the plasma membrane to internal membranes that include the endoplasmic reticulum (ER) and outer mitochondrial membrane. This translocation is associated with cell death. Here we have explored the mechanism of phospho–K-Ras4B toxicity and found that GTP-bound, phosphorylated K-Ras4B associates with inositol trisphosphate receptors on the ER in a Bcl-xL–dependent fashion and, in so doing, blocks the ability of Bcl-xL to potentiate the InsP3 regulated flux of calcium from ER to mitochondria that is required for efficient respiration, inhibition of autophagy, and cell survival. Thus, we have identified inositol trisphosphate receptors as unique effectors of K-Ras4B that antagonize the prosurvival signals of other K-Ras effectors.

Source: Phosphorylated K-Ras limits cell survival by blocking Bcl-xL sensitization of inositol trisphosphate receptors
 
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EMBO J. 1999 Nov 15;18(22):6349-61.
Apoptosis driven by IP(3)-linked mitochondrial calcium signals.
Szalai G1, Krishnamurthy R, Hajnóczky G.
Author information
1 Department of Pathology, Anatomy and Cell Biology, Room 253 JAH, Thomas Jefferson University, Philadelphia, PA 19107, USA.
Abstract
Increases of mitochondrial matrix [Ca(2+)] ([Ca(2+)](m)) evoked by calcium mobilizing agonists play a fundamental role in the physiological control of cellular energy metabolism. Here, we report that apoptotic stimuli induce a switch in mitochondrial calcium signalling at the beginning of the apoptotic process by facilitating Ca(2+)-induced opening of the mitochondrial permeability transition pore (PTP). Thus [Ca(2+)](m) signals evoked by addition of large Ca(2+) pulses or, unexpectedly, by IP(3)-mediated cytosolic [Ca(2+)] spikes trigger mitochondrial permeability transition and, in turn, cytochrome c release. IP(3)-induced opening of PTP is dependent on a privileged Ca(2+) signal transmission from IP(3) receptors to mitochondria. After the decay of Ca(2+) spikes, resealing of PTP occurs allowing mitochondrial metabolism to recover, whereas activation of caspases is triggered by cytochrome c released to the cytosol. This organization provides an efficient mechanism to establish caspase activation while mitochondrial metabolism is maintained to meet ATP requirements of apoptotic cell death.

Source: Apoptosis driven by IP(3)-linked mitochondrial calcium signals. - PubMed - NCBI
Apoptosis driven by IP3‐linked mitochondrial calcium signals
 
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OK @Travis , I have a bottle of Cell Forte IP-6 Inositol staring at me. Should I take it?
Just would like to make a note, I saw a Cell Forte IP6 product which contained Cat's Claw (and Stevia) as well, and Cat's Claw can cause cancer through telomeres, see quote by Haidut:

Potentially yes, as most MAO inhibitors become unselective in higher doses. Also, astragalus* is dangerous as it increases telomere length and can drive cancer growth.
*He mentioned Astragalus but according to the source below, Cat's Claw can lengthen telomeres too:

"Cat’s claw protects and lengthens telomeres. A small in vitro study showed that cells treated with cat’s claw extract lengthened telomeres and extended the lifespan of cells by 201%.[7]"


Source: https://www.truemd.com/anti-aging-wellness/ingest-nutrients-that-increase-telomere-length/

Soure on telomeres length and increased cancer risk: https://www.sciencedaily.com/releases/2017/04/170403083123.htm

Just looking out.
 
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Gene. 2016 Apr 25;581(1):1-13. doi: 10.1016/j.gene.2015.12.046. Epub 2015 Dec 28.
Mg(++) requirement for MtHK binding, and Mg(++) stabilization of mitochondrial membranes via activation of MtHK & MtCK and promotion of mitochondrial permeability transition pore closure: A hypothesis on mechanisms underlying Mg(++)'s antioxidant and cytoprotective effects.
Golshani-Hebroni S.
Abstract
Evidence points to magnesium's antioxidant, anti-necrotic, and anti-apoptotic effects in cardio- and neuroprotection. With magnesium being involved in over 300 biochemical reactions, the mechanisms underlying its cytoprotective and antioxidant effects have remained elusive. The profound anti-apoptotic, anabolic, and antioxidant effects of mitochondrion bound hexokinase (MtHk), and the anti-apoptotic, anti-necrotic, and antioxidant functions of mitochondrial creatine kinase (MtCK) have been established over the past few decades. As powerful regulators of the mitochondrial permeability transition pore (PTP), MtHK and MtCK promote anti-apoptosis and anti-necrosis by stabilizing mitochondrial outer and inner membranes. In this article, it is proposed that magnesium is essentially and directly involved in mitochondrial membrane stabilization via (i) Mg(++) ion requirement for the binding of mitochondrial hexokinase (ii) Mg(++)'s allosteric activation of mitochondrial bound hexokinase, and stimulation of mitochondrial bound creatine kinase activities, and (iii) Mg(++) inhibition of PTP opening by Ca(++) ions. These effects of Mg(++) ions are indirectly supplanted by the stimulatory effect of magnesium on the Akt kinase survival pathway. The "Magnesium/Calcium Yin Yang Hypothesis" proposes here that because of the antagonistic effects of Ca(++) and Mg(++) ions in the presence of high Ca(++) ion concentration at MtHK, MtCK, and PTP, magnesium supplementation may provide cytoprotective effects in the treatment of some degenerative diseases and cytopathies with high intracellular [Ca(++)]/ [Mg(++)] ratio at these sites, whether of genetic, developmental, drug induced, ischemic, immune based, toxic, or infectious etiology.

Source: Mg(++) requirement for MtHK binding, and Mg(++) stabilization of mitochondrial membranes via activation of MtHK & MtCK and promotion of mitochondri... - PubMed - NCBI
 
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Webinar by Dr. Shamsuddin on IP6 + inositol, in English. November 2017. Intro is in Dutch. The audio has sound problems, but tends to get less frequent later in the video. Dr. Shamsuddin presents scientific evidence for the anticancer, antiosteoporosis, anti-inflammatory, cardioprotective, blood thinning, cholesterol lowering, antioxidant, atherosclerotic protective and antifibrotic effects of IP6 + inositol:



Also scientific data is shown how it prevents kidney stones and helps diabetes through multiple ways, including regulating insulin secretion by pancreatic beta cells. And how it is shown to be safe when it comes to serum minerals in a large animal study, something IP6 is often criticized for.

Also what is shown is that according to a study, humans become deficient in IP6 if they consume an IP6-poor diet for as little as 2 weeks. To obtain adequate IP6 levels, more than 10 days are needed if IP6 is supplied by the food, or 4 hours by supplements.

Dr. Shamsuddin also tells how he himself has been taking 2 capsules of IP6 + Inositol for 17.5 years as a preventative. He also refers to the Bantu population in South-Africa, who eat 0.5kg of corn on average daily. When they move to Johannesburg (the urban area) the first generation will still maintain its dietary habit of eating corn, and still have low incidence of cancer and other disease. However, the second generation will adapt to the modern western diet, and will have the same disease rates as the white Afrikaners. At the end he answers questions by attendees of the seminar.

He refers to himself as a mitochondriac, mentions Dr. Otto Warburg, and says he always believed that something in the mitochondria plays a key role in cancer formation.

One of the things he mentioned at the end of the seminar, is that some patients who have cancer decide to stop taking it for a while, go on a drug holiday (can't really understand what he says here) and then their cancer comes back, and it is almost as if the cancer takes vengeance on the patient, and the cancer is uncontrollable at that point.

This last thing is quite worrisome, actually. I hope someone knows what Dr. Shamsuddin ment with that!
 
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Receptors Channels. 1993;1(1):11-24.
Widespread expression of inositol 1,4,5-trisphosphate receptor type 1 gene (Insp3r1) in the mouse central nervous system.
Furuichi T1, Simon-Chazottes D, Fujino I, Yamada N, Hasegawa M, Miyawaki A, Yoshikawa S, Guénet JL, Mikoshiba K.
Author information
1
Division of Behavior and Neurobiology, National Institute for Basic Biology, Okazaki, Japan.
Abstract
The expression of inositol 1,4,5-trisphosphate receptor type 1 (InsP3R1) in the mouse central nervous system (CNS) was studied by in situ hybridization. The receptor mRNAs were widely localized throughout the CNS, predominantly in the olfactory tubercle, cerebral cortex, CA1 pyramidal cell layer of the hippocampus, caudate putamen, and cerebellar Purkinje cells, where phosphoinositide turnover is known to be stimulated by various neurotransmitter receptors. In the most abundantly expressing Purkinje cells, InsP3R1 mRNA appeared to be translocated to the distal dendrites, since a strong hybridization density was observed in the molecular layer of the cerebellum. InsP3R protein is known to form tetrameric receptor-channel complex. Our preliminary hybridization data using probes for three distinct InsP3R subtypes showed preferential expression of InsP3R1 in many parts of the CNS. The expression of other receptor subtypes (InsP3R2 and InsP3R3) is less efficient, suggesting that a homotetramer formed of InsP3R1 subtype may play a central part in InsP3/Ca2+ signalling in the neuronal function, whereas a homotetramer of other subtypes and a possible heterotetramer among subtypes may be involved in differential InsP3/Ca2+ signalling. The chromosomal localization of the gene coding for InsP3R1 was confirmed on chromosome 6 but was found to be genetically independent of the Lurcher (Lc) mutation.

Source: Widespread expression of inositol 1,4,5-trisphosphate receptor type 1 gene (Insp3r1) in the mouse central nervous system. - PubMed - NCBI

So there exists a specific protein, indisputably a membrane pore large enough to admit calcium yet not specifically; this will allow passage of both larger and smaller ions:

'The IP₃ receptor, when activated, can conduct all four alkaline earth cations with conductances in the order of Ba²⁺ > Sr²⁺ > Ca²⁺ > Mg²⁺.' ―Yoshida

While true that this particular membrane pore binds inositol triphosphate (KD ≈ 2–100·nM) it also binds ATP, calcineurin, FKBP12, and some variants will bind calmodulin.

'As shown in Fig. 1, the domain contains putative binding sites for various modulators of the channel such as ATP, Ca²⁺, calmodulin, FK506 binding protein 12 (FKBP 12)...' ―Yoshida

'A specific binding site for ATP was detected in purified type 1 IP₃ receptor subunit (65), and two consensus sequences for ATP binding site were found in its amino acid sequence (24, 25), both located in the coupling domain of the receptor.' ―Yoshida

Yutaka Yoshida isn't a physicist, and doesn't even attempt to describe how calcium actually enters the cell; review articles never prove anything, and its common for their authors to be overly-equitable and to not express strong beliefs (lest it turn into a theoretical article). Yet he does cite another review article in his section marked 'IV. Function of IP₃ receptor.'

Bezprozvanny, I. "The inositol 1,4,5-trisphosphate (InsP₃) receptor." The Journal of membrane biology (1995)

'The ability of Mg²⁺ ions to carry substantial currents through these channels is especially striking when the very high hydration energy and extremely slow substitution rate of water molecules in the inner hydration shell of Mg²⁺ ions (Hille, 1992) is taken into consideration. One possible explanation of this observation is that when Mg²⁺ ions pass through the selectivity filters of both intracellular Ca²⁺ channels they are able to keep the inner shell of water molecules. This suggestion implies that the narrowest portion of the channel pore should be at least 10·Å for both channels. An even larger estimate of the pore size (40·Å) was obtained for the RyR (Lindsay et al., 1991) based on the ability of large organic cations like Tris⁺ and TEA⁺ to permeate through these channels.' ―Bezprozvanny

'It could be concluded from the studies of InsP₃R (Bezprozvanny & Ehrlich, 1994) and RyR (Lindsay et al., 1991; Tinker & Williams, 1992) permeation that both channels are rather nonspecific cation selective channels, permeable to Ca²⁺ and monovalent cations.' ―Bezprozvanny
This is a nonspecific channel, and the author seems to doubt the existence of selective calcium channels:

'As an aside, it follows from this discussion that if plasma membrane InsP₃-gated Ca²⁺-selective channels do exist (Kuno & Gardner, 1987; Fujimoto et al., 1992) they must be much more selective for divalent cations than intracellular InsP₃R.' ―Bezprozvanny
The author notes the affinity ATP has for this membrane pore, and also confirms the ionic series of permissivity:

'These authors came to the conclusion that ATP was a necessary cofactor for the activation of what was then a hypothetical InsP₃R (Smith et al., 1985). The role of ATP as an allosteric activator of the InsP₃R was proposed later based on experiments with receptor that was purified and reconstituted into liposomes (Ferris et al., 1990). It was found that 10 gM ATP or nonhydrolyzable ATP analogues dramatically potentiated InsP₃-mediated Ca²⁺ flux into vesicles containing purified InsP₃R. The existence of a specific ATP-binding site on the InsP₃R was also demonstrated in the same report (Ferris et al., 1990).' ―Bezprozvanny

'All four alkaline earth cations tested were able to pass through the InsP₃R with single channel conductances that fall in the sequence Ba > Sr > Ca > Mg. The same order of conductances was reported for the RyR (Tinker & Williams, 1992) although the absolute values of the single channel conductance are approximately twice as large for the RyR.' ―Bezprozvanny
Based on these considerations the inositol triphosphate receptor could just as easily be called the 'ATP barium pore,' the 'membrane FKBP12 receptor,' or the 'membrane nonselective ion channel.' Adenosine triphosphate (ATP) is known for it's propensity for complexing magnesium, which can also pass through the pore, yet will chelate calcium in its absence. The physical forces responsible for determining how much Ca²⁺ enters and exits through this pore had gone unexplained by both of these authors, and no mention of the fact that inositol phosphates chelate calcium. The closest thing to a physical explanation had been the mention of the luminal 'glutamate ring,' an amino acid often post-translationally modified to γ-carboxyglutamate—a calcium chelator and how vitamin K ultimately exerts its calcemic functions.

I would guess that this is simple a pore having no directionality, allowing ions either in or out just the same; it's complete nonspecificity towards both the 'activating' ligand and the ions permitted is freely admitted, and even a simple dialysis bag will allow unidirectional ion flow if a chelator is placed on one side (i.e. EDTA, ATP, IP₃). An experimental study I would like to see conducted would a comparison of Ca²⁺ flux after the addition of inositol triphosphate, adenosine triphosphate, ethylene diamine tetra-acetate, and simple pyrophosphate. I would think any membrane study that makes claims about the ability of a protein to actually cause unidirectional ion flow would need something akin to an 'EDTA dialysis model' serving as a control: Only then can you eliminate underlying dialysis forces from the equation, if they are not significant, or subtract them from the 'receptor-driven flux' if the are; that could prove its ontological status, but so far I have seen nothing to indicate that its anything more than merely an ion pore.
 
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So there exists a specific protein, indisputably a membrane pore large enough to admit calcium yet not specifically; this will allow passage of both larger and smaller ions:

'The IP₃ receptor, when activated, can conduct all four alkaline earth cations with conductances in the order of Ba²⁺ > Sr²⁺ > Ca²⁺ > Mg²⁺.' ―Yoshida

While true that this particular membrane pore binds inositol triphosphate (KD ≈ 2–100·nM) it also binds ATP, calcineurin, FKBP12, and some variants will bind calmodulin.

'As shown in Fig. 1, the domain contains putative binding sites for various modulators of the channel such as ATP, Ca²⁺, calmodulin, FK506 binding protein 12 (FKBP 12)...' ―Yoshida

'A specific binding site for ATP was detected in purified type 1 IP₃ receptor subunit (65), and two consensus sequences for ATP binding site were found in its amino acid sequence (24, 25), both located in the coupling domain of the receptor.' ―Yoshida

Yutaka Yoshida isn't a physicist, and doesn't even attempt to describe how calcium actually enters the cell; review articles never prove anything, and its common for their authors to be overly-equitable and to not express strong beliefs (lest it turn into a theoretical article). Yet he does cite another review article in his section marked 'IV. Function of IP₃ receptor.'

Bezprozvanny, I. "The inositol 1,4,5-trisphosphate (InsP₃) receptor." The Journal of membrane biology (1995)

'The ability of Mg²⁺ ions to carry substantial currents through these channels is especially striking when the very high hydration energy and extremely slow substitution rate of water molecules in the inner hydration shell of Mg²⁺ ions (Hille, 1992) is taken into consideration. One possible explanation of this observation is that when Mg²⁺ ions pass through the selectivity filters of both intracellular Ca²⁺ channels they are able to keep the inner shell of water molecules. This suggestion implies that the narrowest portion of the channel pore should be at least 10·Å for both channels. An even larger estimate of the pore size (40·Å) was obtained for the RyR (Lindsay et al., 1991) based on the ability of large organic cations like Tris⁺ and TEA⁺ to permeate through these channels.' ―Bezprozvanny

'It could be concluded from the studies of InsP₃R (Bezprozvanny & Ehrlich, 1994) and RyR (Lindsay et al., 1991; Tinker & Williams, 1992) permeation that both channels are rather nonspecific cation selective channels, permeable to Ca²⁺ and monovalent cations.' ―Bezprozvanny
This is a nonspecific channel, and the author seems to doubt the existence of selective calcium channels:

'As an aside, it follows from this discussion that if plasma membrane InsP₃-gated Ca²⁺-selective channels do exist (Kuno & Gardner, 1987; Fujimoto et al., 1992) they must be much more selective for divalent cations than intracellular InsP₃R.' ―Bezprozvanny
The author notes the affinity ATP has for this membrane pore, and also confirms the ionic series of permissivity:

'These authors came to the conclusion that ATP was a necessary cofactor for the activation of what was then a hypothetical InsP₃R (Smith et al., 1985). The role of ATP as an allosteric activator of the InsP₃R was proposed later based on experiments with receptor that was purified and reconstituted into liposomes (Ferris et al., 1990). It was found that 10 gM ATP or nonhydrolyzable ATP analogues dramatically potentiated InsP₃-mediated Ca²⁺ flux into vesicles containing purified InsP₃R. The existence of a specific ATP-binding site on the InsP₃R was also demonstrated in the same report (Ferris et al., 1990).' ―Bezprozvanny

'All four alkaline earth cations tested were able to pass through the InsP₃R with single channel conductances that fall in the sequence Ba > Sr > Ca > Mg. The same order of conductances was reported for the RyR (Tinker & Williams, 1992) although the absolute values of the single channel conductance are approximately twice as large for the RyR.' ―Bezprozvanny
Based on these considerations the inositol triphosphate receptor could just as easily be called the 'ATP barium pore,' the 'membrane FKBP12 receptor,' or the 'membrane nonselective ion channel.' Adenosine triphosphate (ATP) is known for it's propensity for complexing magnesium, which can also pass through the pore, yet will chelate calcium in its absence. The physical forces responsible for determining how much Ca²⁺ enters and exits through this pore had gone unexplained by both of these authors, and no mention of the fact that inositol phosphates chelate calcium. The closest thing to a physical explanation had been the mention of the luminal 'glutamate ring,' an amino acid often post-translationally modified to γ-carboxyglutamate—a calcium chelator and how vitamin K ultimately exerts its calcemic functions.

I would guess that this is simple a pore having no directionality, allowing ions either in or out just the same; it's complete nonspecificity towards both the 'activating' ligand and the ions permitted is freely admitted, and even a simple dialysis bag will allow unidirectional ion flow if a chelator is placed on one side (i.e. EDTA, ATP, IP₃). An experimental study I would like to see conducted would a comparison of Ca²⁺ flux after the addition of inositol triphosphate, adenosine triphosphate, ethylene diamine tetra-acetate, and simple pyrophosphate. I would think any membrane study that makes claims about the ability of a protein to actually cause unidirectional ion flow would need something akin to an 'EDTA dialysis model' serving as a control: Only then can you eliminate underlying dialysis forces from the equation, if they are not significant, or subtract them from the 'receptor-driven flux' if the are; that could prove its ontological status, but so far I have seen nothing to indicate that its anything more than merely an ion pore.
Receptors Channels. 1994;2(1):9-22.
Cloning and characterization of human type 2 and type 3 inositol 1,4,5-trisphosphate receptors.
Yamamoto-Hino M1, Sugiyama T, Hikichi K, Mattei MG, Hasegawa K, Sekine S, Sakurada K, Miyawaki A, Furuichi T, Hasegawa M, et al.
Author information
1 Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., Japan.
Abstract
We have cloned cDNAs coding for human type 2 and type 3 and part of type 1 inositol 1,4,5-trisphosphate receptors (IP3Rs). The complete nucleotide sequences for type 2 and type 3 receptors were determined and the pharmacological properties of the latter were characterized. Human type 2 and type 3 IP3Rs are 2701 amino acids and 2671 amino acids long, respectively, and have significant sequence homologies as well as structural similarities including the six membrane-spanning regions near the C-termini when compared with the rat or mouse counterpart. COS-7 cells transfected with human type 3 IP3R showed characteristic inositol 1,4,5-trisphosphate (IP3)-binding properties with Kd values of 28.8 nM. The order of potency of competition with IP3 was Ins(1,4,5)P3 (IP3) > Ins(2,4,5)P3 > Ins(1,3,4,5)P4 > Ins(1,2,3,4,5,6)P6. Type 2 and type 3 IP3Rs were mapped to human chromosomes 12p11 and 6p21, respectively, by in situ hybridization. cDNA cloning of the human IP3Rs allowed us to identify the types of the receptor expressed in various human hematopoietic and lymphoma cell lines. The type 3 receptor was present in all of cell lines tested, while the type 1 or 2 receptor was expressed in only particular cell types. The differential expression of the IP3R types could confer the cell-specific regulation on the IP3/Ca2+ signalling.

Cloning and characterization of human type 2 and type 3 inositol 1,4,5-trisphosphate receptors. - PubMed - NCBI

Characterization of Inositol Trisphosphate Receptor Binding in Brain REGULATION BY pH AND CALCIUM*
(Received for publication, April 7, 1987)
Paul F. Worley, Jay M. BarabanS, Surachai Supattapone, Virginia S. Wilson, and Solomon H. Snyder8 From the Departments of Neuroscience, Neurology, Phnrmncology, and Molecular Sciences, and Psychiatry and Behavioral Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205

Inositol 1,4,5-trisphosphate is an intracellular second messenger, produced upon stimulation of the phosphoinositide system, capable of mobilizing calcium from intracellular stores. We have recently identified high levels of specific binding sites for inositol 1,4,5- trisphosphate in brain membranes (Worley, P. F., Baraban, J. M., Colvin, J. s., and Snyder, s. H. (1987) Nature 325, 159-161) and have now further characterized these sites. In cerebellar membranes, inositol 1,4,5-trisphosphate binding sites are abundant (20 pmol/mg protein) and display high affinity and selectivity for inositol 1,4,5-trisphosphate (KO = 40 nM), whereas other inositol phosphates such as inositol 1,3,4,5-tetrakisphosphate (Ki = 10 p~) and inositol 1,4-bisphosphate (Ki 3 10 p~) exhibit much lower affinity for this site. Submicromolar concentrations of calcium strongly inhibit inositol 1,4,5-trisphosphate binding (ICao = 300 nM). A sharp increase in binding occurs at slightly alkaline pH. These results suggest that actions of inositol 1,4,5-trisphosphate are regulated by physiological alterations in intracellular pH and calcium concentrations.

http://www.jbc.org/content/262/25/12132.full.pdf

Solubilization, Purification, and Characterization of an Inositol Trisphosphate Receptor*
(Received for publication, July 30, 1987)
Surachai Supattapone, Paul F. Worley, Jay M. BarabanS, and Solomon H. Snyder8
From the Departments of Neuroscience, Pharmacology and Molecular Sciences, Psychiatry and Behavioral Sciences, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205

Inositol 1,4,5-trisphosphate is a second messenger of the phosphoinositide system which can mobilize calcium from intracellular stores. Rat cerebellum is an abundant source of a receptor for inositol 1,4,5-trisphosphate (Worley, P. F., Baraban, J. M., Supattapone, S., Wilson, V. S., and Snyder, S. H. (1987) J. Biol. Chem. 262, 12132-12136). In this study we have solubilized and purified this receptor to apparent homogeneity from rat cerebellum. Crude membrane, detergent-solubilized, and purified receptor preparations display similar selectivity for inositol 1,4,5-trisphosphate over other inositol phosphates. The purified receptor is globular with a Stokes’ radius of approximately 10 nm. Electrophoretic analysis reveals one protein band with an M, of 260,000. While binding is reversibly inhibited by 300 I1M calcium in particulate fractions and detergent-solubilized membranes, the purified protein is not inhibited by calcium concentrations up to 1.5 mM. Inhibition by calcium is reconstituted by addition of detergent-solubilized cerebellar membranes, but not by the cytosolic fraction of cerebellum.

http://www.jbc.org/content/263/3/1530.full.pdf

So what you're saying is Travis, it's basically one big international conspiracy to sell IP6?
 

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