• Microbiology and Biotechnology Letters
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• v.18 no.3
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• pp.233-238
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• 1990

The mechanism of intracellular accumulation of cadmium in a cadmium-ion tolerant yeast, Hansenula ammala B-7, which is an extreme cadmium tolerant strain and has the ability to take up a large amount of cadmium was investigated. The amounts of cadmium taken up by the scalded yeast cells were 2 to 3 times more than the value of the living cells. The living Hansenula anomala B-7 cells adsorbed 74% of cadmium taken up onto the other layer of the cells and 26% of it accumulated inside the cells. But the scalded cells adsorbed 98.3% of cadmium taken up and accumulated 1.7% of it inside the cells. A cadmium uptake and its accumulation were accelerated up to 162.3% and 275.4% by Triton X-100 in the living cells, respectively. Whereas in the scalded cell cadmium uptake was not affected by Triton X-100. Furthermore the cadmium uptake and its accumulation were strongly inhibited by metabolic inhibitors like 2,4-dinitrophenol, sodium azide and potassium cyanide in the living cells, but in the scalded cells cadmium uptake was not affected by metabolic inhibitors. These results suggested that the intracellular accumulation of cadmium by the cadmium-tolerant Hansenula anomala B-7 cells was apparently dependent of biological activity, and also gave evidence of the existance of energy-dependent system.

• The Korean Journal of Physiology and Pharmacology
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• v.3 no.3
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• pp.357-364
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• 1999

We investigated the role of $Ca^{2+}$ and protein kinases/phosphatases in the stimulatory effect of insulin on glucose transport. In isolated rat adipocytes, the simple omission of $CaCl_2$ from the incubation medium significantly reduced, but did not abolish, insulin-stimulated 2-deoxy glucose (2-DG) uptake. Pre-loading adipocytes with intracellular $Ca^{2+}$ chelator, 5,5'-dimethyl bis (o-aminophenoxy)ethane-N,N,N'N' tetraacetic acetoxymethyl ester (5,5'-dimethyl BAPTA/AM) completely blocked the stimulation. Insulin raised intracellular $Ca^{2+}$ concentration $([Ca^{2+}]_i)$ about 1.7 times the basal level of $72{\pm}5$ nM, and 5,5'-dimethyl BAPTA/AM kept it constant at the basal level. This correlation between insulin-induced increases in 2-DG uptake and $[Ca^{2+}]_i$ indicates that the elevation of $[Ca^{2+}]_i$ may be prerequisite for the stimulation of glucose transport. Studies with inhibitors (ML-9, KN-62, cyclosporin A) of $Ca^{2+}-calmodulin$ dependent protein kinases/phosphatases also indicate an involvement of intracellular $Ca^{2+}.$ Additional studies with okadaic acid and calyculin A, protein phosphatase-1 (PP-1) and 2A (PP-2A) inhibitors, indicate an involvement of PP-1 in insulin action on 2-DG uptake. These results indicate an involvement of $Ca^{2+}-dependent$ signaling pathway in insulin action on glucose transport.

• Animal cells and systems
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• v.2 no.2
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• pp.233-238
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• 1998

Certain basic characteristics of choline uptake in nerve terminals were studied with synaptosomes from rat hippocampus. Synaptosomal $[^3H]$-choline uptake was clarified as specific and high affinity by low Km value(2.2 uM), Na+-dependency and high sensitivity to hemicholinium-3, a competitive inhibitor of choline uptake. Choline uptake into synaptosomes was linearlys related to Na+ concentration and membrane potential. Extracellular Ca2+ modulated the choline uptake, but probably not through increase of intracellular $Ca^{2+}$, because this modulation was not affected the by high $K^+$-depolarization. EGTA (2mM) added for $Ca^{2+}$-free condition had a peculiar effect of decreasing choline uptake. These results suggest that Ca2+ may play an important role in regulating the metabolism of acetylcholine in the nerve terminals directly through the increase of acetylcholine release.

• The Korean Journal of Pharmacology
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• v.8 no.1
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• pp.67-75
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• 1972

Many studies on the mechanism of the inotropic action of cardiac glycosides have shown the possible intimate relationship between the mobilization of intracellular calcium and inotropic effect. Evidence obtained from recent studies suggests that cardiac glycosides may increase the intracellular $Ca^{++}$ concentration through the release of this ion from cellular or intracellular membrane. It seemed imperative to study the effect of ouabain on the interaction between mitochondria and $Ca^{++}$, because mitochondria are known to have a rather powerful $Ca^{++}$ pump mechanism which may have an important role on the regulation of intracellular $Ca^{++}$ concentration. The present investigations was made into the effect of ouabain on $Ca^{++}$ untake of mitochondria in the presence of ATP and its dependence on $K^+$ and $Na^+$ in the medium. The results are summarized as follows: 1. The rate of rise in the turbidity of superprecipitation was solely influenced by ionic strength of the medium, not by the species of ion, i.e. $Na^+$ or $K^+$. The higher ionic strength suppressed and the lower enhanced the rate of superprecipitation respectively. 2. No effect of ouabain was found on the rate of superprecipitation. 3. Mitochondria depressed the rate of superpretipitation, and the depressed rate of superprecipitation by mitochondria was reversed by ouabain, and the degree of this reversal was almost identical in $Na^+$ and $K^+$ medium. 4. $Ca^{++}$ uptake of mitochondria was inhibited by ouabain in the presence of ATP and the degree of inhibition showed the dose-response manner in terms of concentration of ouabain. 5. In the absence of ATP, mitochondria took or the $Ca^{++}$ in initial period but released it later. Such uptake and release of $Ca^{++}$ was not influenced by ouabain. 6. It is suggested that intracellular calcium mobilization by ouabain through the action upon the mitochondria was due to inhibition on ATP-dependent $Ca^{++}$ uptake by this agent, not to the action upon so called binding.

• Journal of Microbiology
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• v.33 no.4
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• pp.302-306
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• 1995

E. coli was tested for their ability to uptake cadmium intracellularly, and the effect of zinc and calcium on cadmium toxicity to E. coli was observed. In addition, the effect of zinc and calcium on the uptake of cadimium was also studied. This study showed that living E. coli cells took up more cadmium than the dead cells. E. coli in the log phase uptake cadimiumm more actively than E. coli in the stationary phase. These results suggested that there may be metabolic reactions or compounds which encourage the uptake of cadimium. This study also showed that cadimium was sequestered by cell components of which molecular weight is about 30,000. Adding of zinc and calcium chloride reduced cadmium toxicity in E. coli and encouraged intracellular uptake by E coli. However adding of heavy metal solutions helped the microorganisms to adsorb more cadmium. Extremely high or low concentrations of zinc, however, did not affect cell viability.

• Archives of Pharmacal Research
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• v.25 no.5
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• pp.718-723
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• 2002

CKD-602 (7-[2-(N-isopropylamino)ethyl]-(20S)-camptothecin) is a recently-developed synthetic camptothecin analogue and currently under clinical development by Chong Kun Dang Pharm (Seoul, Korea). CKD-602 showed potent topoisomerase inhibitory activity in vitro and broad antitumor activity against various human tumor cells in vitro and in vivo in animal models. This study describes the pharmacodynamics of the immediate and delayed cytotoxicity induced by CKD-602 in a human colorectal adenocarcinoma cell line, HT-29, and its intracellular drug accumulation by HPLC. The present study was designed to address whether the higher activity of CKD-602 with prolonged exposure is due to delayed exhibition of cytotoxicity and/or an accumulation of anti proliferative effect on continuous drug exposure. The drug uptake study was performed to determine whether the delayed cytotoxicity is due to a slow drug accumulation in cells. CKD-602 produced a cytotoxicity that was exhibited immediately after treatment (immediate effect) and after treatment had been terminated (delayed effect). Both the immediate and delayed effects of CKD-602 showed a time dependent decrease in 4IC_{50}$ values. Drug uptake was biphasic and the second equilibrium level was obtained as early as at 24hr, indicating that the cumulative and delayed antitumor effects of CKD-602 were not due to slow drug uptake. On the other hand, CKD-602 treatment was sufficient to induce delayed cytotoxicity after 4hr, however, longer treatment (＞24hr) enhanced its cytotoxicity due to the intracellular accumulation of the drug, which requires 24hr to reach maximum equilibrium concentration. In addition, $C^n$$\times$T=h analysis (n=0.481) indicated that increased exposure times may contribute more to the overall antitumor activity of CKD-602 than drug concentration. Additional studies to determine the details of the intracellular uptake kinetics (e.g., concentration dependency and retention studies) are needed in order to identify the optimal treatment schedules for the successful clinical development of CKD-602.

• Archives of Pharmacal Research
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• v.19 no.2
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• pp.132-136
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• 1996

Exogenously applied oxygen free radical generating agent, pyrogallol, highly elevated extracellular glutamate accumulation and augmented N-methyl-D-aspartate (NMDA)-induced glutamate accumulation in cerebellar granule neuronal cells, but did not in astrocytes. Superoxide dismutase remarkably decreased the pyrogallol-induced glutamate accumulation, but either NMDA or kainate antagonists did not. In addition, pyrogallol did not affect the NMDAinduced intracellular calcium elevation. Pyrogallol partially blocked glutamate uptake into astrocytes. These results suggest that oxygen free radicals elevate extracellular glutamate accumulation by stimulating the release of glutamate as well as blocking the glutamate uptake.

• Molecules and Cells
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• v.23 no.1
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• pp.11-16
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• 2007

N,N-dimethyl-D-erythro-sphingosine (DMS) is an N-methyl derivative of sphingosine and an inhibitor of protein kinase C (PKC) and sphingosine kinase (SK). In the present study, we examined the effects of DMS on intracellular $Ca^{2+}$ concentration, pH, and glutamate uptake in human 1321N1 astrocytes. DMS increased intracellular $Ca^{2+}$ concentration and cytosolic pH in a concentration-dependent manner. Pretreatment of the cells with the $G_{i/o}$ protein inhibitor PTX and the PLC inhibitor U73122 had no obvious effect. However, removal of extracellular $Ca^{2+}$ with the $Ca^{2+}$ chelator EGTA or depletion of intracellular $Ca^{2+}$ stores with thapsigargin impeded the DMS-induced increase of intracellular $Ca^{2+}$ concentration. Pretreatment of cells with $NH_4Cl$ or monensin reduced the DMS-induced $Ca^{2+}$ increase. However, inhibition of the DMS-induced $Ca^{2+}$ increase with BAPTA did not influence the DMS-induced pH increase. DMS also inhibited glutamate uptake by the 1321N1 astrocytes in a concentration-dependent manner. It also increased intracellular $Ca^{2+}$ and pH in PC12 neuronal cells. Our observations on the effects of DMS on 1321N1 astrocytes and PC12 neuronal cells point to a physiological role of DMS in the brain.

• Toxicological Research
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• v.11 no.2
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• pp.199-203
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• 1995

The focus of this study was to investigate that cellular parameters and glucose uptake might be altered by extracellular calcium and starvation. Addition of 1 mM $Ca^{++}$ to hepatocytes (equalling to the free calcium concentration of blood) significantly increased intracellular $Na^+$ and decreased $Na^+$ & LDH leakage. This pertains to the hepatocytes of control rats as well as those of rats fasted for 24 and 48. hr. These effects might be come from the membrane-stabilizing effects of calcium. But calcium had no effects on cell volumes, superoxide-formation and glucose uptake. Actually hepatocytes of starved rats showed changes in several cellular parameters. Starvation increased LDH leakage, glucose uptake and the total concentration of $Na^+$ and $Na^+$ whereas it markedly decreased cell volumes. Since total tonicity remained unchanged, intracellular $Na^+$ and $Na^+$ could contribute to a higher share of total osmolarity in starvation. Starvation increased the cytoplasmic pH because $R-NH^{3+}$ions and their corresponding counterions disappeared. This increase may be related to suppress the protonization of amino groups in proteins. Starvation decreased hepatic glycogen, a major compound that affects cytosolic volume of hepatocytes. The data indicate that starvation increases the glucose transport activity. The possible molecular basis will be discussed.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.967-971
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• 2004

The kinetics of sulfite uptake were examined in a wild-type laboratory strain of Saccharomyces cerevisiae to determine if carrier-mediated sulfite uptake involved a proton symport, as previous studies on sulfite uptake have suggested both an active process and facilitated diffusion. Accumulation of intracellular sulfite was initially rapid and linear up to 50 sec. Uptake was saturable at final concentrations equal to or greater than 3 mM sulfite, and increased 2-fold in the presence of 2% glucose. Uptake was significantly reduced in cells pretreated with 100-500 $\mu$M carbonyl cyanide mchlorophenylhydrazone (CCCP) or 2,4-dinitrophenol (DNP), both of which dissipate proton gradients. Uptake was also significantly inhibited in the presence of 1 mM arsenate, an inhibitor of ATP synthesis. Extracellular alkalization was observed in cells incubated with 1-2 mM sulfite in a weak tartrate buffer at pH 3.5 and 4.5. These findings suggest that the bisulfite ion, $HSO_3^-$, an anionic form of sulfite, is taken up by a carrier-mediated proton symport. A met16 sull sul2 mutant, impaired in both sulfite formation and sulfate uptake, was found able to grow on a medium with sulfite as the sole Sulfur source, indicating that the sulfate transporters Sul1p and Sul2p are not required for sulfite uptake.