• Journal of Korean Neurosurgical Society
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• v.45 no.4
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• pp.231-235
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• 2009

Objective : While many factors contribute to aging, changes in calcium homeostasis and calcium related neuronal processes are likely to be important. High intracellular calcium is toxic to cells and alterations in calcium homeostasis are associated with changes in calcium-binding proteins, which confine free $Ca^{2+}$. We therefore assayed the expression of the calcium binding proteins calretinin and calbindin in the central auditory nervous system of rats. Methods : Using antibodies to calretinin and calbindin, we assayed their expression in the cochlear nucleus, superior olivary nucleus, inferior colliculus, medial geniculate body and auditory cortex of young (4 months old) and aged (24 months old) rats. Results : Calretinin and calbindin staining intensity in neurons of the cochlear nucleus was significantly higher in aged than in young rats (p<0.05) The number and staining intensity of calretinin-positive neurons in the inferior colliculus, and of calbindin-positive neurons in the superior olivary nucleus were greater in aged than in young rats (p<0.05). Conclusion : These results suggest that auditory processing is altered during aging, which may be due to increased intracellular $Ca^{2+}$ concentration, consequently leading to increased immunoreactivity toward calcium-binding proteins.

• The Korean Journal of Physiology and Pharmacology
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• v.1 no.5
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• pp.565-572
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• 1997

In the freshly isolated rat nephron, the effect of endothelin-1, -2 and -3 (ET-1, -2 and -3) on cytosolic free calcium concentration ($[Ca^{2+}]_i$) was determined using the fluorescent indicator Fura-2/AM. $[Ca^{2+}]_i$ increase was investigated in 9 parts of the single nephron including glomerulus (Glm), $S_1,\;S_2,\;S_3$, cortical and medullary thick ascending limb and cortical (CCT) and outer medullary collecting tubule (OMCT). Endothelins increased $[Ca^{2+}]_i$ in Glm (ET-1; $127{\pm}17%$, ET-2; $93{\pm}5%$, ET-3; $169{\pm}17%$), CCT (ET-1; $30{\pm}6%$, ET-2; $38{\pm}19%$, ET-3; $158{\pm}18%$) and OMCT (ET-1; $197{\pm}11%$, ET-2; $195{\pm}11%$, ET-3; $215{\pm}37%$) at 10-7 M. In OMCT, ET-1 and ET-2 increased $[Ca^{2+}]_i$ in a dose-dependent manner ($10^{-10}{\sim}10^{-6}$ M). To the contrary, ET-3-induced $[Ca^{2+}]_i$ rise was begun from $10^{-12}$ M. BQ-123Na, an antagonist of ETA receptor, at $10^{-4}$ M inhibited about 30% of $[Ca^{2+}]_i$ rise induced by ET-1 and -3. Binding experiments using $[^{125}I]ET-3$ showed the existence of $ET_B$ receptor in OMCT. This binding was replaced by ET-1, ET-2 or ET-3 by the almost same degree but not by angiotensin II or vasopressin.

• The Korean Journal of Physiology and Pharmacology
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• v.9 no.1
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• pp.55-62
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• 2005

Very little research has been carried out on safflower seed for the prevention and treatment of the bone deficiency diseases, including osteoporosis, which are supported by scientific evidences. In the present study, $3{\mu}l$ of 0.1% dried crude extract or $2{\mu}l$ of 0.1% dried aqueous fraction were shown to significantly accelerate the rate of differentiation of osteoblast. Also, the crude extract and aqueous fraction increased the $[Ca^{2+}]_i$ of the cultured osteoblast cells: $3{\mu}l$ of 0.1% dried crude extract and $2{\mu}l$ of 0.1% dried aqueous fraction significantly increased the $[Ca^{2+}]_i$ of the cultured osteoblast cells ($8{\times}10^{-4}$) to the extent that it deserves a considerable attention. Furthermore, the crude extract and aqueous fraction increased the $[Ca^{2+}]_i$ of the cultured osteoblast cells, and $300{\mu}M$ $Cd^{2+}$, specific calcium channel blocker, completely blocked the increase. Therefore, the increased $[Ca^{2+}]_i$ of the cultured osteoblast cells by safflower seed component continued to activate calcium channel.

• Journal of Life Science
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• v.19 no.7
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• pp.892-898
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• 2009

MIF is a progesterone analogue and is known as a potent progesterone antagonist. Although MIF has been known to inhibit prostate cancer cell growth, its molecular mechanisms are not yet clear. In the present study, when the cells were treated for 2-4 days with 5-40 $\mu$M of MIF, the growth and viability of LNCaP cells were significantly decreased in a dose- and time-dependent manner. When the cells, cultivated in a normal 2 mM calcium concentration medium, were treated with 15 $\mu$M MIF for 1 day, the intracellular calcium level increased by 26% compared to the control. Similar results were also found in cells located in the calcium-free reaction buffer, indicating that MIF induced the increase of intracellular Ca$^{2+}$ levels, regardless of the presence of calcium in the surrounding medium. In the cells treated with various concentrations of MIF, the intracellular calcium levels increased in a dose dependent manner. Cells treated with MIF revealed typical early apoptotic signs, i.e., chromosome condensation and nuclei fragmentation. In cells treated with 40 11M MIF, Bcl-2 decreased to 19% of the control. The expression of Bax increased to almost 2 fold of the control. These results demonstrated very clearly that MIF treatment blocks the expression of Bcl-2 but stimulates the expression of Bax. According to the results of the present investigation, the apoptotic mechanism of MIF is triggered by intracellular modulation.

• YAKHAK HOEJI
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• v.43 no.6
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• pp.809-817
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• 1999

The effect of 2-(4-cyanophenyl)amino-1,4-naphthalenedione-3-pyridinium perchlorate (PQ5) on pla-telet aggregation and its action mechanisms were investigated with rat platelet. PQ5 inhibited the platelet aggregation induced by collagen ($6{\;}{\mu\textrm{g}}/ml$), thrombin (0.4 U/ml) and A23187 ($3{\mu}M$) in concentration-dependent manner with $IC_{50}$ values of 5.50, 25.89 and $37.12{\;}{\mu}M$, respectively. PQ5 also significantly reduced the thromboxane $A_2$ (TXA2) formation in a concentration dependent manner. The collagen-induced arachidonic acid (AA) release in [-3H]-AA incorporated platelet, an indication of the phospholipase $A_2$ activity, was decreased by PQ5 pretreatment PQ5 significantly inhibited the activity of thormboxane synthase only at high concentration ($100{\mu}M$), but did not affect the cyclooxygenase activity at all. Collagen-induced ATP release was significantly reduced by PQ5. Calcium-induced platelet aggregation experiment suggests that the elevation of intracellular free $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) by collagen stimulation is decreased by the pretreatment of PQ5, which is due to the inhibition of calcium release from intracellular store and influx from outside of the cell. PQ5 did not showed the effect of anticoagulation as prothrombin time (PT) or activated partial thromboplastin time (APTT). Form these results, it is suggested that PQ5 exerts its antiplatelet activity through the inhibition of the intracellular $Ca^{2+}$ mobilization and the decrease of the $TXA_2$ synthesis.

• YAKHAK HOEJI
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• v.43 no.6
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• pp.802-808
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• 1999

Vibrio vulnificus cytolysin has been incriminated as one of the important virulence determinants in V. vulnificus infection. In the present study, the effects of Vibrio vulnificus cytolysin on platelets were examined. Vibrio vulnificus cytolysin induced platelet aggregation and increased intracellular calcium concentration ($[Ca^{2+}]_i$) of rat platelets. These effects were abolished in $Ca^{2+}-free$ buffer (2 mM EGTA). Cytolysin also potentiated ADP-and collagen-induced platelet aggregation. Lanthanum (2 mM) inhibited cytolysin-diduced platelet aggregation. However, another $Ca^{2+}$ channel blockers, verapamil ($20{\;}{\mu}M$) or mefenamic acid ($20{\;}{\mu}M$) did not block cytolysin-induced platelet aggregation. Osmotic protectants, sucrose (50 mM) and raffinose (50 nM) suppressed platelet aggregation by 35.9% and 63.4%, respectively. V. vulnificus cytolysin increased membrane conductances of platelet membranes. These results suggest that cytolysin-induced platelet aggregation is mediated via lanthanum sensitive-calcium influx which resulted from the pore formation by V. vulnificus cytolysin.

• BMB Reports
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• v.31 no.5
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• pp.468-474
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• 1998

Membrane depolarization in PC12 cells induces calcium influx via an L-type voltage-sensitive calcium channel (L-VSCC) and increases intracellular free calcium, which leads to tyrosine phosphorylation of epidermal growth factor (EGF) receptor and the associated adaptor protein, She. This activated EGF receptor complex then can activate mitogen-activated protein (MAP) kinase, as in nerve growth factor (NGF) receptor activation. In the present study, we investigated the role of EGF receptor in the signaling pathway initiated by membrane depolarization of PC12 cells. Prolonged membrane depolarization induced phosphorylation of extracellular signal-regulated kinase (ERK) within 1 min in undifferentiated PC12 cells. Pretreatment of PC12 cells with the calcium chelator EGTA abolished depolarization-stimulated ERK phosphorylation, but NGF-induced phosphorylation of ERK was not affected. The chronic treatment of phorbol ester, which down-regulated the activity of protein kinase C (PKC), did not affect the phosphorylation of ERK upon depolarization. In the presence of an inhibitor of EGF receptor, neither depolarization nor calcium ionophore increased the level of ERK phosphorylation. These data imply that the EGF receptor is functionally necessary to activate ERK and neurite outgrowth in response to the prolonged depolarization in PC12 cells, and also that PKC is apparently not involved in this signaling pathway.

• Korean Journal of Microbiology
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• v.31 no.1
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• pp.79-84
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• 1993

Infection of Chinook Salmon Embryo (CHSE) cells with IPNV resulted in a significant decrease in intracellular free calcium concentration ([$Ca^{2+}$]i) compared to mock-infected cells. The degree of the decrease in [Ca$^{2+}$]i was dependent on the amount of input virus, and treatment of IPNV-infected CHSE cells with metabolic inhibitors such as cyloheximide cordycepin partially reversed the decrease in [$Ca^{2+}$]i in IPNV-infected cells. Inactiation of PINV with UV also abolished IPNV-induced decrease in [$Ca^{2+}$]i. These data suggest an active role of IPNV in the decrease of [Ca$^{2+}$]i in the infected CHSE cells. The importance of the decrease in [$Ca^{2}$i] could be supported by the finding that the production of IPNV plaques increased in the cells treated with verapamil, a calcium influex blocker, and by lowering the concentration of extracellular calcium. Decreased production of IPNV plaques was observed by elevating the extracellular calcium. Thus, it is suggested that IPNV induced a decreased in [$Ca^{2+}$]i and the decrease in [$Ca^{2+}$]i may plan an importat role in efficient replication of IPNV.ation of IPNV.

• The Korean Journal of Physiology and Pharmacology
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• v.3 no.4
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• pp.439-445
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• 1999

This study is to investigate the mechanism of inhibitory effect of imipramine on the calcium utilization in single cells isolated from canine detrusor. 2 mm thick smooth muscle chops were incubated in 0.12% collagenase solution at $36^{circ}C,$ and aerated with 95% $O_2/5%\;CO_2,$ and then cell suspension was examined. Acetylcholine (ACh) evoked a concentration-dependent contraction of the isolated detrusor cells in normal physiologic salt solution (PSS), and the ACh-induced contraction was significantly inhibited by imipramine. In $Ca^{2+}-free$ PSS, ACh-induced contraction was less than those in normal PSS and it was not affected by the pretreatment with imipramine. $Ca^{2+}-induced$ contraction in $Ca^{2+}-free$ PSS was supressed by imipramine, but addition of A 23187, a calcium ionophore, overcomed the inhibitory effect of imipramine. High potassium-depolarization (40 mM KCl) evoked cell contraction, which was inhibited by imipramine. Caffeine, a releasing agent of the stored $Ca^{2+}$ from sarcoplasmic reticulum, evoked a contraction of the cells that was not blocked by the pretreatment with imipramine. These results suggest that imipramine inhibits the influx of calcium in the detrusor cells through both the receptor-operated- and voltage-gated-calcium channels, but does not affect the release of calcium from intracellular storage site.

• Asian-Australasian Journal of Animal Sciences
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• v.15 no.7
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• pp.949-956
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• 2002

There are several physiological and pharmacological evidences indicating that opening of voltage dependent $Ca^{2+}$ channels play a critical role in induction of acrosome reaction in mammalian sperm. We determined the intracellular free $Ca^{2+}$ concentration in ejaculated goat sperm using a fluorescent, $Ca^{2+}$-specific probe, Fura2/AM, after the suspension of sperm in KRB medium, capable of sustaining capacitation and the acrosome reaction. We used nifedipine, D-600 and diltiazem, the $Ca^{2+}$ channel antagonists belonging to the classes of dihydropyridines, phenylalkylamines and benzothiazepines, to investigate the possibility that L-type voltage gated $Ca^{2+}$ channels play a role in the progesterone-stimulated exocytotic response. Progesterone promoted a rise in intracellular $Ca^{2+}$ in goat sperm and addition of nifedipine (100 nM) just prior to progesterone induction, significantly inhibited both intracellular $Ca^{2+}$ rise and exocytosis suggesting that $Ca^{2+}$ channels are involved in the process. However, the intracellular $Ca^{2+}$ increase during the process of capacitation was not affected with the addition of nifedipine suggesting a role of focal channel for $Ca^{2+}$ during capacitation. Studies using monensin and nigericin, two monovalent cation ionophores showed that an influx of $Na^+$ also may play a role in the opening of $Ca^{2+}$ channels. These results strongly suggests that the entry of $Ca^{2+}$ channels with characteristics similar to those of L-type, voltage-sensitive $Ca^{2+}$ channels found in cardiac and skeletal muscle, is a crucial step in the sequence of events leading to progesterone induced acrosome reaction in goat sperm.