Blockade of Ca3 calcium channels and induction of G/G cell cycle arrest in colon cancer cells by gossypol.

Background And Purpose: Gastrointestinal tumours overexpress voltage-gated calcium (Ca3) channels (Ca3.1, 3.2 and 3.

Heterogeneity of the endoplasmic reticulum Ca store determines colocalization with mitochondria.

Contact sites between the endoplasmic reticulum (ER) and mitochondria play a pivotal role in cell signaling, and the interaction between these organelles is dynamic and finely regulated. We have studied the role of ER Ca concentration ([Ca]) in modulating this association in HeLa and HEK293 cells and human fibroblasts. According to Manders' coefficient, ER-mitochondria colocalization varied depending on the ER marker; it was the highest with ER-Tracker and the lowest with ER Ca indicators (Mag-Fluo-4, erGAP3, and G-CEPIA1er) in both HeLa cells and human fibroblasts.

PKC Inhibits Sec61 Translocon-Mediated Sarcoplasmic Reticulum Ca Leak in Smooth Muscle Cells.

PKC inhibitors stimulate Ca release from internal stores in diverse cell types. Our data indicate that this action cannot be explained by an increased agonist-induced IP production or an overloaded SR Ca pool in smooth muscle cells from guinea pig urinary bladder. The incubation of these cells with three different PKC inhibitors, such as Go6976, Go6983, and BIM 1, resulted in a higher SR Ca leak revealed by inhibition of the SERCA pump with thapsigargin.

Aldosterone-Induced Sarco/Endoplasmic Reticulum Ca Pump Upregulation Counterbalances Ca1.2-Mediated Ca Influx in Mesenteric Arteries.

In mesenteric arteries (MAs), aldosterone (ALDO) binds to the endogenous mineralocorticoid receptor (MR) and increases the expression of the voltage-gated L-type Ca1.2 channel, an essential ion channel for vascular contraction, sarcoplasmic reticulum (SR) Ca store refilling, and Ca spark generation. In mesenteric artery smooth muscle cells (MASMCs), Ca influx through Ca1.

The Increased Expression of Regulator of G-Protein Signaling 2 (RGS2) Inhibits Insulin-Induced Akt Phosphorylation and Is Associated with Uncontrolled Glycemia in Patients with Type 2 Diabetes.

Experimental evidence in mice models has demonstrated that a high regulator of G-protein signaling 2 (RSG2) protein levels precede an insulin resistance state. In the same context, a diet rich in saturated fatty acids induces an increase in RGS2 protein expression, which has been associated with decreased basal metabolism in mice; however, the above has not yet been analyzed in humans. For this reason, in the present study, we examined the association between RGS2 expression and insulin resistance state.

Caspase-3 Activation Correlates With the Initial Mitochondrial Membrane Depolarization in Neonatal Cerebellar Granule Neurons.

In this study we evaluated the effect of the reduction in the endoplasmic reticulum calcium concentration ([Ca]), changes in the cytoplasmic calcium concentration ([Ca]), alteration of the mitochondrial membrane potential, and the ER stress in the activation of caspase-3 in neonatal cerebellar granule cells (CGN). The cells were loaded with Fura-2 to detect changes in the [Ca] and with Mag-fluo-4 to measure variations in the [Ca] or with TMRE to follow modifications in the mitochondrial membrane potential in response to five different inducers of CGN cell death. These inducers were staurosporine, thapsigargin, tunicamycin, nifedipine and plasma membrane repolarization by switching culture medium from 25 mM KCl (K25) to 5 mM KCl (K5).

Sarco-Endoplasmic Reticulum Calcium Release Model Based on Changes in the Luminal Calcium Content.

The sarcoplasmic/endoplasmic reticulum (SR/ER) is the main intracellular calcium (Ca) pool in muscle and non-muscle eukaryotic cells, respectively. The reticulum accumulates Ca against its electrochemical gradient by the action of sarco/endoplasmic reticulum calcium ATPases (SERCA pumps), and the capacity of this Ca store is increased by the presence of Ca binding proteins in the lumen of the reticulum. A diversity of physical and chemical signals, activate the main Ca release channels, i.

Polymerization of sarcoplasmic-reticulum calcium-binding proteins might explain observed reticulum kinetics-on-demand behavior.

Reported experimental results, in which transient elevations of sarcoplasmic calcium levels are induced by caffeine in smooth muscle cells, apparently contradict the principle of mass conservation. The commonly accepted model assumes that the total number of Ca binding sites is fixed. A former work dealing with this problem proved that assuming the presence within the reticulum of calcium sequestering proteins whose total number of calcium binding sites increases as the existent sites get occupied, is enough to explain the above referred counter-intuitive experimental results.

The Trans Golgi Region is a Labile Intracellular Ca Store Sensitive to Emetine.

The Golgi apparatus (GA) is a bona fide Ca store; however, there is a lack of GA-specific Ca mobilizing agents. Here, we report that emetine specifically releases Ca from GA in HeLa and HL-1 atrial myocytes. Additionally, it has become evident that the trans-Golgi is a labile Ca store that requires a continuous source of Ca from either the external milieu or from the ER, to enable it to produce a detectable transient increase in cytosolic Ca.

Differential expression and signaling of the human histamine H receptor isoforms of 445 and 365 amino acids expressed in human neuroblastoma SH-SY5Y cells.

In stably-transfected human neuroblastoma SH-SY5Y cells, we have compared the effect of activating two isoforms of 445 and 365 amino acids of the human histamine H receptor (hHR and hHR) on [S]-GTPγS binding, forskolin-induced cAMP formation, depolarization-induced increase in the intracellular concentration of Ca ions ([Ca]i) and depolarization-evoked [H]-dopamine release. Maximal specific binding (B) of [H]-N-methyl-histamine to cell membranes was 953 ± 204 and 555 ± 140 fmol/mg protein for SH-SY5Y-hHR and SH-SY5Y-hHR cells, respectively, with similar dissociation constants (K, 0.86 nM and 0.

Activation of endoplasmic reticulum calcium leak by 2-APB depends on the luminal calcium concentration.

It has been shown that 2-APB is a nonspecific modulator of ion channel activity, while most of the channels are inhibited by this compound, there are few examples of channels that are activated by 2-APB. Additionally, it has been shown that, 2-APB leads to a reduction in the luminal endoplasmic reticulum Ca level ([Ca]) and we have carried out simultaneous recordings of both [Ca] and the [Ca] in HeLa cell suspensions to assess the mechanism involved in this effect. This approach allowed us to determine that 2-APB induces a reduction in the [Ca] by activating an ER-resident Ca permeable channel more than by inhibiting the activity of SERCA pumps.

Orai3 channel is the 2-APB-induced endoplasmic reticulum calcium leak.

We have studied in HeLa cells the molecular nature of the 2-APB induced ER Ca leak using synthetic Ca indicators that report changes in both the cytoplasmic ([Ca]) and the luminal ER ([Ca]) Ca concentrations. We have tested the hypothesis that Orai channels participate in the 2-APB-induced ER Ca leak that was characterized in the companion paper. The expression of the dominant negative Orai1 E106A mutant, which has been reported to block the activity of all three types of Orai channels, inhibited the effect of 2-APB on the [Ca] but did not decrease the ER Ca leak after thapsigargin (TG).

Increased calcium leak associated with reduced calsequestrin expression in hyperthyroid cardiomyocytes.

Introduction: Calcium (Ca) leak during cardiac diastole is chiefly mediated by intracellular Ca channel/Ryanodine Receptors. Increased diastolic Ca leak has been proposed as the mechanism underlying the appearance of hereditary arrhythmias. However, little is known about alterations in diastolic Ca leak and the specific roles played by key intracellular Ca-handling proteins in hyperthyroidism, a known arrhythmogenic condition.

Histamine H receptor activation stimulates calcium mobilization in a subpopulation of rat striatal neurons in primary culture, but not in synaptosomes.

The histamine H receptor (HR) is abundantly expressed in the Central Nervous System where it regulates several functions pre and postsynaptically. HRs couple to Gα proteins and trigger or modulate several intracellular signaling pathways, including the cAMP/PKA pathway and the opening of N- and P/Q-type voltage-gated Ca channels. In transfected cells, activation of the human HR of 445 amino acids (hHR) results in phospholipase C (PLC) stimulation and release of Ca from intracellular stores.

Palmitic acid but not palmitoleic acid induces insulin resistance in a human endothelial cell line by decreasing SERCA pump expression.

Palmitic acid is a negative regulator of insulin activity. At the molecular level, palmitic acid reduces insulin stimulated Akt Ser473 phosphorylation. Interestingly, we have found that incubation with palmitic acid of human umbilical vein endothelial cells induced a biphasic effect, an initial transient elevation followed by a sustained reduction of SERCA pump protein levels.

Kinetics on Demand Is a Simple Mathematical Solution that Fits Recorded Caffeine-Induced Luminal SR Ca2+ Changes in Smooth Muscle Cells.

The process of Ca2+ release from sarcoplasmic reticulum (SR) comprises 4 phases in smooth muscle cells. Phase 1 is characterized by a large increase of the intracellular Ca2+ concentration ([Ca2+]i) with a minimal reduction of the free luminal SR [Ca2+] ([Ca2+]FSR). Importantly, active SR Ca2+ ATPases (SERCA pumps) are necessary for phase 1 to occur.

Endoplasmic reticulum stress in insulin resistance and diabetes.

The endoplasmic reticulum is the main intracellular Ca(2+) store for Ca(2+) release during cell signaling. There are different strategies to avoid ER Ca(2+) depletion. Release channels utilize first Ca(2+)-bound to proteins and this minimizes the reduction of the free luminal [Ca(2+)].

Calcium signalling in diabetes.

Molecular cascades responsible for Ca(2+) homeostasis and Ca(2+) signalling could be assembled in highly plastic toolkits that define physiological adaptation of cells to the environment and which are intimately involved in all types of cellular pathology. Control over Ca(2+) concentration in different cellular compartments is intimately linked to cell metabolism, because (i) ATP production requires low Ca(2+), (ii) Ca(2+) homeostatic systems consume ATP and (iii) Ca(2+) signals in mitochondria stimulate ATP synthesis being an essential part of excitation-metabolic coupling. The communication between the ER and mitochondria plays an important role in this metabolic fine tuning.

Acidic intracellular Ca(2+) stores and caveolae in Ca(2+) signaling and diabetes.

Acidic Ca(2+) stores, particularly lysosomes, are newly discovered players in the well-orchestrated arena of Ca(2+) signaling and we are at the verge of understanding how lysosomes accumulate Ca(2+) and how they release it in response to different chemical, such as NAADP, and physical signals. Additionally, it is now clear that lysosomes play a key role in autophagy, a process that allows cells to recycle components or to eliminate damaged structures to ensure cellular well-being. Moreover, lysosomes are being unraveled as hubs that coordinate both anabolism via insulin signaling and catabolism via AMPK.

Ryanodine receptors as leak channels.

Ryanodine receptors are Ca(2+) release channels of internal stores. This review focuses on those situations and conditions that transform RyRs from a finely regulated ion channel to an unregulated Ca(2+) leak channel and the pathological consequences of this alteration. In skeletal muscle, mutations in either CaV1.

An intelligent sarco-endoplasmic reticulum Ca2+ store: release and leak channels have differential access to a concealed Ca2+ pool.

Simultaneous recording of cytosolic and sarco-endoplasmic reticulum (SR/ER) luminal free calcium concentrations ([Ca(2+)](i) and [Ca(2+)](L), respectively) supports the notion that release channels (RyRs and IP(3)Rs) use a concealed Ca(2+) source, likely to be associated with intra-SR/ER Ca(2+) binding proteins, whereas SR/ER Ca(2+) leak channels can only access free luminal Ca(2+). We hypothesize that Ca(2+) is trapped by oligomers of luminal Ca(2+)-binding proteins and that the opening of release channels induces the rapid liberation of this "concealed" Ca(2+) source associated with intra-ER Ca(2+) buffers. Our hypothesis may also clarify why SERCA pumps potentiate Ca(2+) release and explain quantal characteristics and refractory states of Ca(2+) release process.

Inhibition of SERCA pumps induces desynchronized RyR activation in overloaded internal Ca2+ stores in smooth muscle cells.

We have previously shown that rapid inhibition of sarcoplasmic reticulum (SR) ATPase (SERCA pumps) decreases the amplitude and rate of rise (synchronization) of caffeine induced-Ca(2+) release without producing a reduction of free luminal SR Ca(2+) level in smooth muscle cells (Gómez-Viquez L, Guerrero-Serna G, García U, Guerrero-Hernández A. Biophys J 85: 370-380, 2003). Our aim was to investigate the role of luminal SR Ca(2+) content in the communication between ryanodine receptors (RyRs) and SERCA pumps.

Variable luminal sarcoplasmic reticulum Ca(2+) buffer capacity in smooth muscle cells.

Sarcoplasmic reticulum contains the internal Ca(2+) store in smooth muscle cells and its lumen appears to be a continuum that lacks diffusion barriers. Accordingly, the free luminal Ca(2+) level is the same all throughout the SR; however, whether the Ca(2+) buffer capacity is the same in all the SR is unknown. We have estimated indirectly the luminal Ca(2+) buffer capacity of the SR by comparing the reduction in SR Ca(2+) levels with the corresponding increase in [Ca(2+)](i) during activation of either IP(3)Rs with carbachol or RyRs with caffeine, in smooth muscle cells from guinea pig urinary bladder.