• The Korean Journal of Physiology and Pharmacology
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• v.3 no.2
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• pp.199-205
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• 1999

Vibrio vulnificus cytolysin caused platelet cytolysis and increased intracellular calcium concentration $([Ca^{2+}]_i)$ of rat platelets in a concentration-dependent manner. In the presence of V. vulnificus cytolysin (3 HU/ml), lactate dehydrogenase (LDH) activity was increased from $1.3{\pm}0.4%$ of control to $64.3{\pm}3.4%$ in platelet suspension buffer. In $Ca^{2+}-free$ platelet suspension buffer, however, V. vulnificus cytolysin did not induce $[Ca^{2+}]_i$ increase and LDH release. Addition of EGTA (2 mM) to suspension buffer after the initial $Ca^{2+}$ influx reversed $[Ca^{2+}]_i$ to the control level. However, a $Ca^{2+}$ channel blocker verapamil $(20\;{\mu}M)$ or mefenamic acid $(20\;{\mu}M)$ did not inhibit V. vulnificus cytolysin-induced $[Ca^{2+}]_i$ increase and LDH release. Divalent cations such as $Co^{2+},\;Cd^{2+}\;or\;Mn^{2+}$ (2 mM each) also did not alter V. vulnificus cytolysin-induced $[Ca^{2+}]_i$ increase and LDH release. V. vulnificus cytolysin (3 HU/ml)-induced calcium influx was completely blocked by lanthanum (2 mM). Lanthanum (2 mM) also completely blocked V. vulnificus cytolysin (3 HU/ml)-induced LDH release. Osmotic protectants such as, raffinose, sucrose or PEG600 (50 mM each) did not inhibit the lytic activity of V. vulnificus cytolysin. In conclusion, lanthanum sensitive $Ca^{2+}$ influx plays a significant role in Vibrio vulnificus cytolysin-induced platelet cytolysis and thrombocytopenia in V. vulnificus infection.

• The Korean Journal of Physiology
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• v.21 no.2
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• pp.191-200
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• 1987

The activation mechanism of the sustained contractions induced by norepinephrine and K-depolarization was studied in renal vascular muscle. Helical strips of arterial muscle were prepared from rabbit renal arteries. All experiments were performed in Tris-buffered Tyrode solution which was aerated with 100% $O_2$ and kept at $35^{\circ}C$. Renal arterial muscles developed a contracture rapidly when exposed to a 40 mM K-Tyrode solution. In the absence of external $Ca^{2+}$, however, no K-contracture appeared. The contracture induced by K-depolarization was abolished by the treatment with $Ca^{2+}-antagonist\;(verapamil)$ or lanthanum $(La^{3+})$. From these results, it is obvious that K-contracture of renal arterial strip required $Ca^{2+}$ in the medium and this contracture was developed by the increased $Ca^{2+}-influx$ due to K-depolarization. Noradrenaline (5 mg/l) induced also a similar sustained contraction rapidly in all strips. Even on the K-contracture and in $Ca^{2+}-free$ Tyrode solution and also in the Tyrode solution pretreated with verapamil or $La^{3+}$, noradrenaline produced a contraction. However, the contraction in $Ca^{2+}-free$ Tyrode solution was not sustained and decreased gradually. The amplitude of noradrenaline-induced contracture was dependent on external $Ca^{2+}$; The contracture increased dose-dependently, but over 3 mM $Ca^{2+}$, decreased. The results of this experiment suggest that K-contracture was developed by an increased $Ca^{2+}-influx$ due to membrane depolarization, while noradrenaline-induced contracture was developed by both transmembrane $Ca^{2+}-influx$ and the mobilizaiton of cellular $Ca^{2+}$

• Animal cells and systems
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• v.7 no.1
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• pp.75-79
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• 2003

Highly purified high performance thin layer chromatography (HPTLC) fractions containing a putative calcium influx factor (CIF) were prepared from the Jurkat cells and Xenopus oocytes in which $Ca^{2+}$ stores were depleted by thapsigargin treatment and from the yeast in which intracellular $Ca^{2+}$ stores were also depleted by genetic means. Microinjection of the fractions has been shown to elicit $Ca^{2+}$ dependent currents in Xenopus oocytes. The nature of the membrane currents evoked by the putative CIF appeared to be carried by chloride ions since the current was blocked by the selective chloride channel blocker 1 mM niflumic acid and its reversal potential was about -24 mV. Injection of the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N, N, N',N'-tetraacetic acid (BAPTA) eradicated the current activities, suggesting the current responses are entirely $Ca^2$-dependent. Moreover, the currents were sensitive to the removal of extracellular calcium, indicating the dependence on calcium entry through the plasma membrane calcium entry channels. CIF activities were insensitive to protease, heat, and acid treatments and to Dische-reaction whereas the activities were sensitive to nucleotide pyrophosphatase and hydrazynolysis. The fraction might have a sugar because it was sensitive to Molisch test and Seliwaniff's resorcinol reaction. From the above results, CIF as a small and stable molecule seems to have pyrimidine, pyrophosphate, and a sugar moiety.oiety.

• Biomedical Science Letters
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• v.15 no.4
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• pp.273-279
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• 2009

Cordycepin (3'-deoxyadenosine) is an adenosine analogue isolated from Cordyceps militaris, and it has been used as an anti-cancer and anti-inflammation ingredient in traditional Chinese medicine. We investigated the effects of cordycepin on human platelet aggregation induced by thapsigargin, and determined the cytosolic free $Ca^{2+}$ levels ($[Ca^{2+}]_i$), an aggregation-stimulating factor. Cordycepin significantly inhibited thapsigargin-induced platelet aggregation. Its inhibitory effect was continually sustained at the maximal aggregation concentration of thapsigargin. The thapsigargin-induced $[Ca^{2+}]_i$ were clearly reduced by cordycepin in the presence of exogenous $CaCl_2$ or extracellular $Ca^{2+}$-chelator (EDTA). These results suggest that cordycepin inhibited thapsigargin-induced $Ca^{2+}$-influx from extracellular domain and thapsigargin-induced $Ca^{2+}$-mobilization from intracellular $Ca^{2+}$ storage. Accordingly, our data demonstrated that cordycepin may have a beneficial effect on platelet aggregation-mediated thrombotic diseases by inhibiting a $[Ca^{2+}]_i$-elevation.

• The Korean Journal of Physiology
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• v.23 no.2
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• pp.329-337
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• 1989

Cardiac muscles show stimulation frequency-dependent tension changes i.e. Bowditch phenomenon and Woodworth phenomenon, the former is an increase of tension with the increase of stimulation frequency, whereas the latter is an increase of tension with a decrease of stimulation frequency. Bowditch phenomenon is seen in the range of frequency 1.0 cps and above, and Woodworth phenomenon below the frequency 1.0 cps in the most of mammalian cardiac atrium. To throw some light on the possible mechanism of both phenomena in rat atrium, influences of drugs affecting $Ca^{2+}$ influx through the plasma membrane $(verapamil,\;La^{3+},\;norepinephrine)$ and $Ca^{2+}$ release from sarcoplasmic reticulum (SR) on frequency-tension curve were studied. The results obtained are summarized as follows: 1) At low temperature $(27.5^{\circ}C)$, both Bowditch and Woodworth phenomenon were demonstrated. But Bowditch phenomenon disappeared at the temperature above $(32.5^{\circ}C)$. 2) At $(27.5^{\circ}C)$, in the presence of verapamil, a $Ca^{2+}$ channel blocker, a time course of change in the frequency-tension was studied. It was found that Bowditch phenomenon was affected before the Woodworth phenomenon, then the former was completely disappeared. At $(32.5^{\circ}C)$, where no Bow-ditch is seen in normal atrial muscle, Bowditch phenomenon was reappeared by an administration of norepinephrine suggesting again that slow inward current of such as $Ca^{2+}$ channel is closely related to Bowditch phenomenon. 3) At $27.5^{\circ}C$, in the presence of $La^{3+}$, although tensions were decreased at all stimulation frequencies, Bowditch and Woodworth phenomenon were still demonstrated. However in the presence of both $La^{3+}$ and verapamil, Bowditch phenomena was disappeared suggesting that $La^{3+}$ is less effective in blocking $Ca^{2+}$ channel than verapamil. 4) At $27.5^{\circ}C$, in the presence of ryanodine, an inhibitor of calcium release from SR, Woodworth phenomenon was disappeared, which was consistent with previous reports of others, suggesting that $Ca^{2+}$ release from SR is closely related to Woodworth phenomenon. From the above findings, it may be concluded that Bowditch phenomenon is dependent on the magnitude of $Ca^{2+}$ influx through slow channel and Woodworth phenomenon is dependent on the amount of $Ca^{2+}$ stored in SR.

• Natural Product Sciences
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• v.12 no.4
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• pp.217-220
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• 2006

The effects of $(1R,9S)-\beta-hydrastine$ hydrochloride (BHSH) on $Ca^{2+}$ transport in rat pheochromocytoma PC12 cells were investigated. In the presence of external $Ca^{2+}$, BHSH at $100{\mu}M$ inhibited $K^+$ (56mM)-induced dopamine release, and $K^+-induced$ $Ca^{2+}$ influx and a sustained rise of $[Ca^{2+}]_i$. In addition, BHSH at 100 f.!M reduced the sustained rise of $[Ca^{2+}]_i$ elicited by 20 mM caffeine, but not by $1{\mu}M$ thapsigargin, in presence of external $Ca^{2+}$. These results suggest that BHSH inhibited $K^+-induced$ dopamine release and $[Ca^{2+}]_i$ influx, and store-operated $Ca^{2+}$ channels activated by caffeine, but not by thapsigargin, in PC12 cells.

• The Korean Journal of Zoology
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• v.39 no.2
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• pp.182-189
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• 1996

It has been known that spermatozoa should obtain their fertilizing ability through capacitation and acrosome reaction, and that in these processes of fertilization, Ca2+ platys an important role for their conjugation. Therefore the present study has been designed in order to clarify the effect of fluctuation of the media Ca2+ level and the intracellular concentration of the spermatozoa on the acrosomes. During the incubation of spermatozoa, a considerable fluctuation in the media Ca2+ level has been observed after the BSA administration and the media concentration of Ca2+. It is deduced that these fluctuation rates may have an effect on the acrosome reaction. The fluctuation of K+ flux has been observed in accordance with the incubation period over time, and it's concentration seems to be closely related with the acrosomal reaction. The respiratory exchange rate (RERI of the spermatozoa is kept more regular in the BSA and Cacl2 administration groups than the non-administration group. Based on the experimental findings, it is possible to deduce a hypothesis from these findings that physiological activities of the acrosome reaction are not functionally related to the media Ca2+ level and the intracellular influx of Ca2+ concentration, although Ca2+ platys an important role as a stimulating factor in the acrosome reaction.

• The Korean Journal of Pharmacology
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• v.32 no.3
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• pp.417-424
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• 1996

The present study was done to examine the involvement of protein kinase C and protein tyrosine kinase in intracellular $Ca^{2+}$ mobilization in C5a-stimulated neutrophils. Although protein kinase C inhibitors, staurosporine and H-7 inhibited intracellular $Ca^{2+}$ release in C5a-stimulated neutrophils, they did not affect $Ca^{2+}$ influx across the plasma membrane and elevation of $[Ca^{2+}]_i$ C5a-induced intracellular $Ca^{2+}$ release and $Ca^{2+}$ influx were inhibited by protein tyrosine kinase inhibitors, genistein and methyl-2,5-dihydroxycinnamate. ADP-evoked elevation of $[Ca^{2+}]_i$ was inhibited by genistein and methyl-2,5-dihydroxycinnamate but was not affectd by staurosporine and H-7. Genistein and methyl-2,5-dihydroxycinnamate reduced the store-regulated $Ca^{2+}$ influx in thapsigargin-treated neutrophils, while the effect of staurosporine and H-7 was not detected. When neutrophils were preincubated wih phorbol 12-myristate 13-acetate, the stimulatory effect of C5a on the elevation of $[Ca^{2+}]_i$ was reduced. These results suggest that protein tyrosine kinase may be involved in control of intracellular $Ca^{2+}$ release and $Ca^{2+}$ influx across the plasma membrane in C5a-activated neutrophils.

• Molecules and Cells
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• v.19 no.2
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• pp.268-278
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• 2005

We investigated the effect of cytosolic and extracellular $Ca^{2+}$ on $Ca^{2+}$ signals in pancreatic acinar cells by measuring $Ca^{2+}$ concentration in the cytosol($[Ca^{2+}]_c$) and in the lumen of the ER($[Ca^{2+}]_{Lu}$). To control buffers and dye in the cytosol, a patch-clamp microelectrode was employed. Acetylcholine released $Ca^{2+}$ mainly from the basolateral ER-rich part of the cell. The rate of $Ca^{2+}$ release from the ER was highly sensitive to the buffering of $[Ca^{2+}]_c$ whereas ER $Ca^{2+}$ refilling was enhanced by supplying free $Ca^{2+}$ to the cytosol with $[Ca^{2+}]_c$ clamped at resting levels with a patch pipette containing 10 mM BAPTA and 2 mM $Ca^{2+}$. Elevation of extracellular $Ca^{2+}$ to 10 mM from 1 mM raised resting $[Ca^{2+}]_c$ slightly and often generated $[Ca^{2+}]_c$ oscillations in single or clustered cells. Although pancreatic acinar cells are reported to have extracellular $Ca^{2+}$-sensing receptors linked to phospholipase C that mobilize $Ca^{2+}$ from the ER, exposure of cells to 10 mM $Ca^{2+}$ did not decrease $[Ca^{2+}]_{Lu}$ but rather raised it. From these findings we conclude that 1) ER $Ca^{2+}$ release is strictly regulated by feedback inhibition of $[Ca^{2+}]_c$, 2) ER $Ca^{2+}$ refilling is determined by the rate of $Ca^{2+}$ influx and occurs mainly in the tiny subplasmalemmal spaces, 3) extracellular $Ca^{2+}$-induced $[Ca^{2+}]_c$ oscillations appear to be triggered not by activation of extracellular $Ca^{2+}$-sensing receptors but by the ER sensitised by elevated $[Ca^{2+}]_c$ and $[Ca^{2+}]_{Lu}$.

• The Korean Journal of Physiology
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• v.20 no.1
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• pp.31-41
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• 1986

The sufficient myoplasmic $Ca^{++}$ to react with the contractile proteins is necessary to induce contraction of a cardiac muscle. These $Ca^{++}$ for the production of muscle contraction are supplied from the three recognized $Ca^{++}$ sources; internal $Ca^{++}$ release via the sarcoplasmic reticulum(SR), $Ca^{++}$ influx through a gated Ca-channel in the membrane as a Isi, and $Ca^{++}$ transport by the mechanism of Na/ca exchange. However, it is still controversial which $Ca^{++}$ sources act as a main contributor for myoplasmic $Ca^{++}$, Therefore, this study was undertaken in order to examine the $Ca^{++}$ sources for the contraction of frog ventricle. There is evidence that the SR is sparse in frog ventricular fibers, and that T-tubules are absent. Isolated ventricular strips of frog, Rana nigromaculata, were used in this experiment. Isometric tension was recorded by force transducer, and membrane potentials of ventricular muscles were measured through the intracellular glass microelectrodes, which were filled with 3M KCI and had resistance of $30{\pm}50M{\Omega}$. All experiments were performed at room temperature in a tris·buffered Ringer solution which was aerated with 100% $O_2$. Isotonic high K, low Na solution was used to induce K-contracture, K-contracture appeared at the concentration of 20 to 30mM-KCI and was potentiated in parallel with the increase in KCI concentration. The contracture had two components: an initial rapid phasic and a subsequent slow tonic contractile responses. Membrane Potentials measured at normal Ringer solution(2.5mM KCI) was -90 to -100 mV, and decreased linearly as the KCI concentration increased; -55mV at 20mM.KCI, -45mV at 30 mM.KCI, -30 mY at 50 mM.KCI, and -12 mV at 100 mM.KCI. K-contracture was evoked firstly at the membrane potential of -45 mV. The contracture was potentiated by the increase of bathing extracellular $Ca^{++}$ concentration. However, in the absence of $Ca^{++}$ the contracture was almost not induced by 50 mM.KCI solution. Caffeine(20mM) in normal Ringer solution, which is known to release $Ca^{++}$ from SR without substantial effects on the $Ca^{++}$ fluxes across the surface membrane, did not affect membrane potential and also not initiate contracture, but the caffeine in 20 mM-KCI Ringer solution produced a contracture. Above results suggest that the main $Ca^{++}$ source for the K·contracture of frog ventricle is $Ca^{++}$ influx through the voltage-dependent Ca-channel, and that in the K-contracture at the concentration of 100 mM-KCI, the mechanism of Na/ca exchange also partly contributs, in addition to the $Ca^{++}$ influx.