• Applied Biological Chemistry
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• v.39 no.3
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• pp.189-194
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• 1996

The effects of scorpion (Leiurus quinquestriatus hebraeus, Lqh) venom were evaluated on the activities of microsomal $Ca^{2+}-ATPase$ and $Ca^{2+}$ release channel prepared from the epithelial cells of pig airway. Whole venom of Lqh $(120\;{\mu}g/ml)$ increased the activity of microsomal $Ca^{2+}-ATPase$ about 32% in the tight-sealed microsomes and about 28% in the Triton X-100-treated or $Ca^{2+}$ ionophore A23187-treated leaky microsomes. Thapsigargin, a specific antagonist of $Ca^{2+}-ATPase$, inhibited 42% of total ATPase activity and also completely blocked the effects of Lqh venom, suggesting that Lqh venom directly activiates the microsomal $Ca^{2+}-ATPase$. In order to determine if Lqh venom increases the microsomal uptake of $^{45}Ca^{2+}$, Lqh venom was added in the uptake medium. The Lqh venom increased microsomal $^{45}Ca^{2+}$ uptake up to ${\sim}20%$ and the increase was only observed when heparin, an antagonist of $InsP_3$ receptor channel, was added in the uptake medium. Lqh venom in the absence of heparin unexpectedly decreased the rate and the amount of $^{45}Ca^{2+}$ uptake. These results were explained by simultaneous increases in $^{45}Ca^{2+}$ release as well as $^{45}Ca^{2+}$ uptake by Lqh venom. Lqh venom itself increased the release of $^{45}Ca^{2+}$ as much as $^{45}Ca^{2+}$ release by $4\;{\mu}m\;InsP_3$, implying that Lqh venom also activates $InsP_3$ receptor, microsomal $Ca^{2+}$ release channel. Based on these results, we suggest that the Lqh venom consists of at least two components; one activates the $InsP_3$ receptor and the other avates the $Ca^{2+}-ATPase$. Currently we a investigating the chemical and electrophysiological properties of the active components of Lqh venom.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.6
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• pp.725-732
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• 1998

In order to elucidate the molecular mechanism of the intracellular $Ca^{2+}$ overload frequently reported from diabetic heart, diabetic rats were induced by the administration of streptozotocin, the membrane vesicles of junctional SR (heavy SR, HSR) were isolated from the ventricular myocytes, and SR $Ca^{2+}$ uptake and SR $Ca^{2+}$ release were measured. The activity of SR $Ca^{2+}-ATPase$ was $562{\pm}14$ nmol/min/mg protein in control heart. The activity was decreased to $413{\pm}30$ nmol/min/mg protein in diabetic heart and it was partially recovered to $485{\pm}18$ nmol/min/mg protein in insulin-treated diabetic heart. A similar pattern was observed in SR $^{45}Ca^{2+}$ uptakes; the specific uptake was the highest in control heart and it was the lowest in diabetic heart. In SR $^{45}Ca^{2+}$ release experiment, the highest release, 45% of SR $^{45}Ca^{2+}$, was observed in control heart. The release of diabetic heart was 20% and it was 30% in insulin-treated diabetic heart. Our results showed that the activities of both SR $Ca^{2+}-ATPase$ and SR $Ca^{2+}$ release channel were decreased in diabetic heart. In order to evaluate how these two factors contribute to SR $Ca^{2+}$ storage, the activity of SR $Ca^{2+}-ATPase$ was measured in the uncoupled leaky vesicles. The uncoupling effect which is able to increase the activity of SR $Ca^{2+}-ATPase$ was observed in control heart; however, no significant increments of SR $Ca^{2+}-ATPase$ activities were measured in both diabetic and insulin-treated diabetic rats. These results represent that the $Ca^{2+}$ storage in SR is significantly depressed and, therefore, $Ca^{2+}-sequestering$ activity of SR may be also depressed in diabetic heart.

• Proceedings of the Korean Biophysical Society Conference
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• 1996.07a
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• pp.39-39
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• 1996

Extracts were prepared from six different poisonous mushrooms in order to identify biologically active natural ligands. The effects of these extracts were examined on the microsomal $^{45}$ Ca$^{2+}$ uptake and $^{45}$ Ca$^{2+}$ release. Five out of six species apparently inhibited the activity of total microsomal ATPases prepared from the epithelial cells of pig airway. (omitted)ted)

• Biomolecules & Therapeutics
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• v.1 no.2
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• pp.171-176
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• 1993

Purpose of the present study was to clarify the role of noradrenergic neural activities in hypothalamus for either triggering or maintaining hypertension in deoxycorticosterone (DOCA)-salt hypertensive rats. Two groups of animals were prepared: 1) normotensive Wistar rats and 2) DOCA-salt induced hypertensive rats. Voltage dependent $^{45}Ca^{++}$ uptake, endogenous norepinephrine release, and the catecholamine content in the hypothalamus of DOCA-salt hypertensive and normotensive Wistar rats were compared. Animals at 4, 6 and 16 week-old of two groups were sacrificed by decapitation and hypothalamus was dissected out. Voltage dependent calcium uptake and norepinephrine release were determined from hypothalamic synaptosomes either in low potassium or high potassium stimulatory condition by using $^{45}Ca^{++}$ isotope and HPLC-ECD technique. Degrees of voltage dependent $^{45}Ca^{++}$ uptake and norepinephrine release in hypothalamic synaptosomes of 16-week-old DOCA-salt hypertensive rats were significantly greater than those of age matched normotensive control rats. The norepinephrine and dopamine contents of hypothalamus were about the same in two groups of animals. These results suggest that the alteration of evoked norepinephrine release related to calcium uptake in hypothalamus may play a role in the maintenance of hypertension in DOCA-salt hypertensive rats.

• Proceedings of the Korean Biophysical Society Conference
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• 1996.07a
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• pp.40-40
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• 1996

Abnormally high $Ca^{2+}$ concentrations have been reported in the cardiac myocytes of diabetic mellitus (DM). In order to elucidate the molecular mechanisms of the intracellular $Ca^{2+}$ overload, the activities of $^{45}$ Ca$^{2+}$ uptake and $^{45}$ Ca$^{2+}$ release were measured from the vesicles of junctional SR (Heavy SR, HSR). (omitted)omitted)

• The korean journal of orthodontics
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• v.22 no.1
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• pp.273-283
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• 1992

To find out the differences between periodontal ligament cells (PDL cells) and gingival fibroblast cells (GFB cells), alkaline phosphatase, a marker enzyme for osteoblast, was used to measure the activities and $^{45}CaCl_2$ isotope was used to find out cellular and release of $^{45}Ca$, a requisite for bone formation,. PDL cells and GFB cells from 1 to 5 passages were also measured in alkaline phosphatase activity assay. By the use of above methods, followings were concluded that the PDL cells and the GFB cells have characteristics that are different from each other. In that PDL cells showed large amount of calcium uptake and large amount of calcium release in initial stage, they seem to possess characteristics which are similar to osteoblast-like cells. 1. The PDL cells, in contrast to the gingival fibroblast, showed exceedingly high alkaline phosphatase activity which was highest at the second passage, decreasing thereon. But gingival fibroblasts cells showed no distinct differences in alkaline phosphatase activity as the passage were elapsed. 2. For both PDL cells and GF cells, the $^{45}Ca$ uptake was greatest at 2 hours period. The PDL cells showed higher measuring than GFB cells through out the whole time period. 3. Whereas the GFB cells showed slow increase of $^{45}Ca$ release as time relapsed, the PDL cells showed rapid increase of $^{45}Ca$ release.

• Journal of Ginseng Research
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• v.18 no.2
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• pp.95-101
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• 1994

Saponin of Panax ginseng (C.A. Meyer) is composed of Protopanaxatriol (PT) and Protopanaxa- diol (PD). We investigated the effects of PT and PD on the contractility and $^{45}Ca$ uptake in the pig coronary artery. Isometric tension in the helical strips and $^{45}Ca$ uptake in the ring strips were measured in the presence or absence of PT and PD. PT and PD did not affect the high K+ (40 mM)-induced contraction but relaxed the ACh-induced contraction in a dose4ependent manner (1~10 mg/dl). The vasorelaxing effect of PT on the ACh-induced contraction was more potent than that of PD. Those relaxations were partially suppressed by the rubbing of endothelium removal. ACh-induced contraction in the $Ca^{2+}$-free Tyrode's solution was suppressed by the pretreatment of PT or PD. Following the depletion of ACh-sensitive intracellular $Ca^{2+}$ pool, ACh-induced contraction was suppressed by the pratreatment of PT or PD. With the pretreatment of PT or PD, $^{45}Ca$ uptake by high K+ (43 mM) was not changed but that by ACh was suppressed in the pig coronary artery. From the above results, we suggested that the vasorelaxing effect of PT and PD of Panax ginseng was due to inhibition of intracellular $Ca^{2+}$ release, inhibition of $Ca^{2+}$ uptake via receptor-operated $Ca^{2+}$ channels and in part a release of vasorelaxing factor from endothelium in pig coronary artery.

• BMB Reports
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• v.34 no.6
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• pp.573-577
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• 2001

The mammalian heart is known to undergo significant mechanical changes during fetal and neonatal development. The objective of this study was to define the ontogeny of the ryanodine receptor/$Ca^{2+}$ release channel and SERCA that play the major roles in excitation-contraction coupling. Whole ventricular homogenates of fetal (F) (19 and 22 days in gestation), postnatal (N) (1 and 7 days postnatal), and adult (A) (5 weeks postnatal) Sprague-Dawley rat hearts were used to study [$^3H$]ryanodine binding and oxalate-supported $^{45}Ca^{2+}$ uptake. For the ryanodine receptor, the major findings were: (1) The ryanodine receptor density, as determined by maximal [$^3H$]ryanodine binding ($B_{max}$), increased 3 fold between the F22 and A periods ($0.26{\pm}0.1$ vs. $0.73{\pm}0.07$ pmoles/mg protein, p<0.01), whereas there was no significant change during the F22 and N1 development phases ($0.26{\pm}0.1$ vs. $0.34{\pm}0.01$). (2) Affinity of the ryanodine receptor to ryanodine did not significantly change, as suggested by the lack of change in the $K_d$ during the development and maturation. For SERCA, changes started early with an increased rate of $Ca^{2+}$ uptake in the fetal periods (F19: $8.1{\pm}1.1$ vs. F22: $19.3{\pm}2.2$ nmoles/g protein/min; p<0.05) and peaked by 7 days (N7) of the postnatal age ($34.9{\pm}2.1$). Thus, we conclude that the quantitative changes occur in the ryanodine receptor during myocardial development. Also, the maturation of the $Ca^{2+}$ uptake appears to start earlier than that of the $Ca^{2+}$ release.

• The Korean Journal of Physiology
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• v.29 no.2
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• pp.269-277
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• 1995

Membrane vesicles were prepared by differential centrifugation from epithelial cells of porcine trachea. Total activity of microsomal ATPases was measured spectrophotometrically by a coupled enzyme assay. The steady-state activity of the enzyme was $329{\pm}10$ nmol/min mg protein. Thapsigargin, a specific antagonist of intracellular $Ca^{2+}-ATPase$, inhibited about 50% of the activity, leaving $178{\pm}18\;nmol/min .mg$ protein (n=6), indicating that the $Ca^{2+}-ATPase$ is one of the major microsomal ATPases. The microsomes used in this study appeared to be tight-sealed vesicles since they showed saturation in $^{45}Ca^{2+}$ uptake experiments. Inositol 1,4,5-trisphosphate $InsP_{3}, 4\;{\mu}M$, an agonist of $InsP_{3}$-sensitive $Ca^{2+}$ release channel ($InsP_{3}$, receptor), and Ca-ionophore A23187 $(10\;{\mu}M)$ induced $^{45}Ca^{2+}$ releases of 20% and 50% of stored $^{45}Ca^{2+}$, respectively. The addition of $(10\;{\mu}M\;InsP_{3}$ also increased the microsomal ATPase activity from $282{\pm}8$ nmol/min mg protein to $334{\pm}21$ nmol/min . mg protein in the intact vesicles. Similar increase in the activity was observed by making microsomes leaky (uncoupling) using the Ca-ionophore A23187. ;$InsP_{3}-induced$ effects were blocked by either thapsigargin or heparin suggesting that: 1) the $InsP_{3}-induced$ increase in ATPase activity is mediated by microsomal $Ca^{2+}-ATPase$, and 2) dissipation of $Ca^{2+}$ gradient across the microsomal membrane is responsible for the $InsP_{3}-induced$ effect. In order to test the dependence of the $Ca^{2+}-ATPase$ activity on the activity of $InsP_{3}-induced$ the activity of ATPases was monitored in various concentrations of free $Ca^{2+}$ using $EGTA-Ca^{2+}$ buffers. The $Ca^{2+}$-dependent biphasic change is the well-known character of $InsP_{3} receptor but not of microsomal $Ca^{2+}-ATPase$ in non-excitable cells; however, the activity of microsomal ATPase appeared biphasic and a maxim진 activity of $397{\pm}36nmol/min\;.mg$ protein was obtained in the solution containing 100 nM free $Ca^{2+}$. Below or above this concentration, the activity of ATPases was lower. These results strongly support a positive correlation of microsomal $Ca^{2+}-ATPase$ to the $InsP_{3}$ receptors in epithelial microsomes.

• Proceedings of the Korean Biophysical Society Conference
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• 2002.06b
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• pp.56-56
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• 2002

Muscle contraction and relaxation are regulated by the sarcoplasmic reticulum (SR)-mediated $Ca^{2＋}$ release and $Ca^{2＋}$ uptake. The SR functions are closely related with the proteins residing in the SR such as ryanodine receptor, $Ca^{2＋}$-ATpase, calsequestrin, triadin and junctin. In an effort to further identify important functional SR proteins, experiments of sucrose-density gradient of SR fractionation, concanavalin A treatment, 2D gel electrophoresis, $^{45}$ Ca$^{2+}$ overlay, Strains-all staining, and peptide finger printing (PFP) were carried out.(omitted)d)