• Archives of Pharmacal Research
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• v.21 no.6
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• pp.664-670
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• 1998

The role of intracellular $Ca^{2+}$, in the regulation of tumor cell proliferation by products of arachidonic acid (AA) metabolism was investigated using U-373 MG human as trocytoma cells. Treatment with nordihydroguaiaretic acid (NDGA), a lipoxygenase (LOX) inhibitor, or caffeic acid (CA), a specific 5-LOX inhibitor, suppressed proliferation of the tumor cells in a dose-dependent manner. However, indomethacin (indo), a cyclooxygenase (COX) inhibitor, did not significantly alter proliferation of the tumor cells. At anti-proliferative concentrations, NDGA and CA significantly inhibited intracellular $Ca^{2+}$ release induced by carbachol, a known intracelluar $Ca^{2+}$ agonist in the tumor cells. Exogenous administration of leukotriene $B_4(LTB_4)$, an AA metabolite of LOX pathway, enhanced proliferation of the tumor cells in a concentration-dependent fashion. In addition, $LTB_4$, induced intracelluar $Ca^{2+}$ release. Intracellular $Ca^{2+}$-inhibitors, such as an intracellular $Ca^{2+}$ chelator (BAPTA) and intracellular $Ca^{2+}$-release inhibitors (dantrolene and TMB-8), significantly blocked the LTB4-induced enhancement of cell proliferation and intracellular $Ca^{2+}$ release. These results suggest that LOX activity may be critical for cell proliferation of the human astrocytoma cells and that intracelluar $Ca^{2+}$ may play a major role in the mechanism of action of LOX.

• YAKHAK HOEJI
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• v.57 no.6
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• pp.388-393
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• 2013

Previously, we have reported that lovastatin, an inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, increased melanin synthesis through intracellular $Ca^{2+}$ release in B16 cells. In this study we investigated the possible involvement of nitric oxide (NO) in the mechanism of lovastatin-induced melanogenesis. Lovastatin elevated NO formation in a dose-dependent manner. Treatment with mevalonate, farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP), precursors of cholesterol, did not significantly alter the lovastatin-induced NO production, suggesting that inhibition of cholesterol metabolism may not be involved in the mechanism of this action of lovastatin. Both NO formation and melanogenesis induced by lovastatin was significantly suppressed by treatment with $N^G$-nitro-L-arginine methyl ester (L-NAME) and 2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethylinidazoline-1-oxyl-3-oxide (cPTIO), an inhibitor of NO synthase and a NO scavenger, respectively. The lovastatin-induced NO production was significantly affected not by EGTA, an extracellular $Ca^{2+}$ chelator, but by an intracellular $Ca^{2+}$ chelator (BAPTA/AM) and intracellular $Ca^{2+}$ release blockers (dantrolene and TMB-8). Taken together, these results suggest that lovastatin may induce melanogenesis through NO formation mediated by intracellular $Ca^{2+}$ release in B16 cells. These results further suggest that lovastatin may be a good candidate for the therapeutic application of various hypopigmentation disorders.

• Biomolecules & Therapeutics
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• v.15 no.3
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• pp.137-144
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• 2007

Although lovastatin, a competitive inhibitor of 3-hydroxy-3-methylglutaryl-coenzyme A (HMGCoA) reductase, has been shown to have anti-cancer actions, the effect on human hepatoma cells was not investigated. Moreover, the exact mechanism of this action is not fully understood. In this study we investigated the mechanism by which lovastatin induces apoptosis using HepG2 human hepatoblastoma cells. Lovastatin induced apoptotic cell death in a dose-dependent manner in the cells, assessed by the flow cytometric analysis. Treatment with mevalonic acid, a precursor of cholesterol, did not significantly suppress the lovastatin-induced apoptosis. Lovastatin induced a rapid and sustained increase in intracellular $Ca^{2+}$ concentration. Treatment with EGTA, an extracellular $Ca^{2+}$ chelator did not significantly alter the lovastatin-induced intracellular $Ca^{2+}$ increase and apoptosis, whereas intracellular $Ca^{2+}$ reduction with BAPTA/AM and intracellular $Ca^{2+}$ release blockers (dantrolene and TMB-8) completely blocked these actions of lovastatin. In addition, the lovastatin-induced apoptosis was significantly reduced by a calpain inhibitor, a broad spectrum caspase inhibitor z-VAD-fmk and inhibitors specific for caspase-9 and caspase-3 (z-LEHD-fmk and z-DEVD-fmk, respectively), but not by an inhibitor specific for caspase-8 (z-IETD-fmk). Collectively, these results suggest that lovastatin induced apoptosis of HepG2 hepatoma cells through intracellular $Ca^{2+}$ release and calpain activation, leading to triggering mitochondrial apoptotic pathway. These results further suggest that lovastatin may be valuable for the therapeutic management of human hepatoma.

• The Journal of Korean Physical Therapy
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• v.19 no.5
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• pp.65-76
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• 2007

Purpose: It is generally accepted that smooth muscle contraction is triggered by intracellular $Ca^{2+}$ ($[Ca^{2+}]_i$) released from intracellular $Ca^{2+}$ stores such as sarcoplasmic teticulum (SR) and from the extracellular space. The increased $[Ca^{2+}]^i$ can phosphorylate the 20,000 dalton myosin light chain $(MLC_{20})$ by activating MLC kinase (MLCK), and this initiates smooth muscle contraction. In addition to the $[Ca^{2+}]_i$MACK-tension pathway, a number of intracellular signal molecules, including mitogen-activated protein kinase (MAPK), protein kinase C (PKC) and others, play important roles in the regulation of smooth muscle contraction. However, the mechanisms regulating contraction of depletion of SR $Ca^{2+}$ in mouse gastric smooth muscle strips is not still clear. Methods: To investigate the rotes of $Ca^{2+}$ influx and SR $Ca^{2+}$ release channel on gastric motility, isometric contraction and $[Ca^{2+}]_i$ were examined in mouse gastric smooth muscle strips. Results: High KCl, ryanodine, an activator of $Ca^{2+-}$induced $Ca^{2+}$ release channel, and cyclopiazonic acid (CPA), an inhibitor of SR $Ca^{2+-}$ATPase evoked a sustained increase in muscle contraction and $[Ca^{2+}]_i$. These increases induced by high KCl, ryanodine, and CPA were partially blocked by application of verapamil ($10{\mu}M$), a L-type $Ca^{2+}$ channel inhibitor. Additionally, in $Ca^{2+-}$free solution (1 mM EGTA), ryanodine and CPA had no effect contraction and $[Ca^{2+}]_i$ in fundic muscle strips. Conclusion: These results that extracellular $Ca^{2+}$ influx and depletion of SR trigger $Ca^{2+}$ influx through verapamil-sensitive $Ca^{2+}$ channel, and extracellular and SR $Ca^{2+}$ store may functionally involve in the subcellular $Ca^{2+}$ mobilization in mouse gastric muscle.

• Korean Journal of Environmental Biology
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• v.21 no.3
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• pp.303-307
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• 2003

Tributyltin (TBT) used world-wide in antifouling paints toy ships is a wide-spread environmental pollutant. At low doses, antiproliferative modes of action have been shown to be involved, whereas at higher doses apoptosis seems to be the mechanism of toxicity in reproductive organs by TBT. In this study, we investigated that the mechanisms underlying apoptosis induced by TBT in R2C cell. Effects of TBT on intracellular $Ca^{2+}$ level and reactive oxygen species (ROS) were investigated in R2C cells by fluorescence detector. TBT significantly induced intracellular $Ca^{2+}$ level in a time-dependent manner. The rise in intracellular $Ca^{2+}$ level was followed by a time-dependent generation of reactive oxygen species (ROS) at the cytosol level. Simultaneously, TBT induced the release of cytochrome c from the mitochondrial membrane into the cytosol. Furthermore, ROS production and the release of cytochrome c were reduced by BAPTA, an intracellular $Ca^{2+}$ chelator, indicating the important role of $Ca^{2+}$ in R2C during these early intracellular events. In addition, Z-DEVD FMB, a caspase -3 inhibitor, decreased apoptosis by TBT. Taken together, the present results indicated that the apoptotic pathway by TBT might start with an increase in intracellular $Ca^{2+}$ level, continues with release of ROS and cytochrome c from mitochondria, activation of caspases, and finally results in DNA fragmentation.

• Journal of Physiology & Pathology in Korean Medicine
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• v.18 no.5
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• pp.1382-1386
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• 2004

This study was performed for the investigation of vasodilatory efficacy and its underlying mechanisms of Samhwangsasim-tang(SST), herbal remedy. SST relaxed vascular strips precontracted with phenylephrine or KCI(51 mM), but the magnitude of relaxation was greater in phenylephrine(PE) induced contraction. The relaxation effects of SST was endothelium-independent. L-NAME, iNOS inhibitor, and methyl en blue(MB), cGMP inhibitor, did not attenuate the relaxation responses of SST. In the absence of extracellular Ca2+, pre-incubation of the aortic rings with SST significantly reduced the contraction by PE, suggesting that the relaxant action of the SST includes inhibition of Ca/sup 2+/ influx and release of Ca/sup 2+/ from intracellular stores (SR). In addition, the cell death was induced by SST in human aortic smooth muscle cells but not that of human umbilical vein endothelial cells. We conclude that in rat thoracic aorta, SST may induce in part vasodilation through inhibition of Ca/sup 2+/ influx and release of Ca/sup 2+/ from intracellular stores.

• The Korean Journal of Physiology and Pharmacology
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• v.14 no.2
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• pp.105-111
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• 2010

Inositol 1,4,5-trisphosphate receptors ($InsP_3Rs$) modulate $Ca^{2+}$ release from intracellular $Ca^{2+}$ store and are extensively expressed in the membrane of endoplasmic/sarcoplasmic reticulum and Golgi. Although caffeine and 2-aminoethoxydiphenyl borate (2-APB) have been widely used to block $InsP_3Rs$, the use of these is limited due to their multiple actions. In the present study, we examined and compared the ability of caffeine and 2-APB as a blocker of $Ca^{2+}$ release from intracellular $Ca^{2+}$ stores and $Ca^{2+}$ entry through store-operated $Ca^{2+}$ (SOC) channel in the mouse pancreatic acinar cell. Caffeine did not block the $Ca^{2+}$ entry, but significantly inhibited carbamylcholine (CCh)-induced $Ca^{2+}$ release. In contrast, 2-APB did not block CCh-induced $Ca^{2+}$ release, but remarkably blocked SOC-mediated $Ca^{2+}$ entry at lower concentrations. In permeabilized acinar cell, caffeine had an inhibitory effect on InsP3-induced $Ca^{2+}$ release, but 2-APB at lower concentration, which effectively blocked $Ca^{2+}$ entry, had no inhibitory action. At higher concentrations, 2-APB has multiple paradoxical effects including inhibition of Ins$P_3$-induced $Ca^{2+}$ release and direct stimulation of $Ca^{2+}$ release. Based on the results, we concluded that caffeine is useful as an inhibitor of $InsP_3R$, and 2-APB at lower concentration is considered a blocker of $Ca^{2+}$ entry through SOC channels in the pancreatic acinar cell.

• Archives of Pharmacal Research
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• v.22 no.1
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• pp.48-54
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• 1999

These studies were designed to examine the differential effect of nitric oxide (NO) and cGMP on glutamate neurotransmission. In primary cultures of rat cerebellar granule cells, the glutamate receptor agonist N-methyl-D-aspartate (NMDA) stimulates the elevation of intracellular calcium concentration ($[Ca^{2+}]_i$), the release of glutamate, the synthesis of NO and an increase of cGMP. Although NO has been shown to stimulate guanylyl cyclase, it is unclear yet whether NO alters the NMDA-induced glutamate release and ${[Ca^{2+}]}_i$ elevation. We showed that the NO synthase inhibitor, NG-monomethyl-L-arginine (NMMA), partially prevented the NMDA-induced release of glutamate and elevation of ${[Ca^{2+}]}_i$ and completely blocked the elevation of cGMP. These effects of NO on glutamate release and [Ca2+]i elevation were unlikely to be secondary to cGMP as the cGMP analogue, dibutyryl cGMP (dBcGMP), did not suppress the effects of NMDA. Rather, dBcGMP slightly augmented the NMDA-induced elevation of ${[Ca^{2+}]}_i$ with no change in the basal level of glutamate or ${[Ca^{2+}]}_i$. The extracellular NO scavenger hydroxocobalamine prevented the NMDA-induced release of glutamate providing indirect evidence that the effect of NO may act on the NMDA receptor. These results suggest that low concentration of NO has a role in maintaining the NMDA receptor activation in a cGMP-independent manner.

• Korean Journal of Veterinary Research
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• v.38 no.4
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• pp.712-719
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• 1998

Thyroid function is mainly regulated through cAMP and phophatidylinositol, and it is well known that TSH-stimulated thyroxine ($T_4$) release is inhibited by catecholamine from mouse thyroids via the ${\alpha}_1$-adrenoceptor stimulation. Previous study has established that the inhibition of $T_4$ release by ${\alpha}_1$-adrenoceptor stimulation results in activated protein kinase C (PKC). The purpose of this study was to determine if ion transport systems are involved in the inhibition of $T_4$ release elicited by ${\alpha}_1$-adrenergic agonist in mouse thyroids. TSH-, IBMX- and cAMP analogue-stimulated $T_4$ release were significantly inhibited by methoxamine, R59022 (diacylglycerol kinase inhibitor), and MDL (adenylate cyclase inhibitor). TSH-stimulated $T_4$ release could be inhibited by Bay K 8644 and cyclopiazoic acid, but not by verapamil and tetrodotoxin. The addition of nifedipine ($Ca^{2+}$ channel blocker), tetrodotoxin and lidocaine ($Na^+$ channel blockers), but not amiloride (EIPA) and ryanodine, completely blocked the inhibitory effects of methoxamine on $T_4$ release. TSH-stimulated $T_4$ release was also inhibited by benzamil ($Na^+-Ca^{2+}$ exchange inhibitor). TSH-, IBMX- and cAMP-stimulated $T_4$ release were inhibited by methoxamine or R59022, these effects were reversed by nifedipine. but not by verapamil. Furthermore, nifedipine reversed the inhibitory effects of benzamil and R59022 on TSH-stimulated $T_4$ release. These data suggest that the observed ${\alpha}_1$-adrenoceptor-mediated inhibition of $T_4$ release in mouse thyroids is the result of an increase in intracellular $Na^+$ or $Ca^{2+}$ effected via activation of fast $Na^+$ or nifedipine-sensitive $Ca^{2+}$ channels, and that $Na^+-Ca^{2+}$ exchange may play an important role in reducing thyroid hormone by increasing intracellular $Ca^{2+}$.

• Journal of Chest Surgery
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• v.25 no.4
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• pp.347-355
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• 1992

The precise mechanism of the Excitation-Contraction Coupling is still uncertain. But the concept that Ca2+ induced Ca2+ release [CICR] from the Ryanodine receptor in the sarcoplasmic reticulum [foot structure] may play a major role in E-C coupling has been widely accepted since 1970`s. It is believed that increased cytosolic Ca2+ followed by CICR is main contributor for E-C coupling of striated muscle. Resulting phenomena of ischemic /post-reperfusion myocyte is increased cytosolic Ca2+, even to the absence of Ca2+ in reperfusate. So intracellular inhibitor to CICR might prevent the ischemic and reperfusion damage of myocardial cells. The relatively purified foot protein, especially heavy sarcoplasmic reticulum rich, of the skeletal muscle was incorporated into the black lipid bilayer [Phosphatidyl ethanolamine: Phosphatidyl serine=l: 1]. Under the steady state of membrane potential [+20 mV], ionic current through Ryanodine receptor was measured with Cs+ as charge carrier. In the cis chamber [Cytoplasmic side], Mg2+ strongly inhibited CICR of Ryanodine receptor[Kd=6.2 nM]. In conclusion, naturally existing intracellular free Mg2+ can inhibit CICR from intracellular Ca2+ reservior [heavy SR]. So post-ischemic or post-reperfusing myocardium could be preserved using additional free Mg2+ in cardioplegic solution or reperfusate, otherwise the optimal concentration is undetermined.