• Journal of Ginseng Research
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• v.20 no.3
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• pp.274-283
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• 1996

In this study, the effects of red ginseng components [ginsenosides (total saponins and $Rg_1$) on the function of ryanodine receptor (RyR) -$Ca^{2+}$ release channel complex protein (named as RyR or $Ca^{2+}$ channel), a membrane protein in sarcoplasmic reticulum (SR) of rabbit skeletal muscle were examined at the SR vesicle's level and the molecular levels with Chaps-solubilized and purified $Ca^{2+}$ channel protein and with reconstituted proteoliposomes by dialysis. The results were as follows. 1. The binding of ryanodine known as inhibitor of muscle contraction to the RyR was decreased at the whole range of concentration ($10^2$~$10^7$%) by these two ginseng components. In heavy SR vesicles, Chaps-solubilized and purified $Ca^{2+}$ channel protein, and reconstituted vesicles, its maximal inhibition by total saponins was shown at the concentration of $10^3$, $10^3$%, and $10^5$% respectively, and by gin- senoside $Rg_1}$) each was $10^3$%, $10^3$%, and $10^4$%. 2. The release of $Ca^{2+}$ ion through $Ca^{2+}$ channel in heavy SR vesicles and reconstituted proteoliposomes was increased as a whole by these two ginseng components, and particularly maximal release by both of them was shown at the range of $10^4$~$10^6$%. These results were seemed to be caused by conformational change of $Ca^{2+}$ release channel protein (RyR) by red ginseng components [ginsenosides (total saponins and $Rg_1}$).

• Proceedings of the Korean Biophysical Society Conference
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• 2001.06a
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• pp.46-46
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• 2001

Evidence has suggested that an anion channel blocker, 4, 4'-diisothiocyanatostilbene-2, 2' disulfonic acid (DIDS) could trigger Ca release from skeletal sarcoplasmic reticulum (SR) by binding to a 30 kDa SR protein. Since the high molecular weight $Ca^{2＋}$ release channel (CRC)/ryanodine receptor (RyR) is the main SR protein that conducts $Ca^{2＋}$ efflux in skeletal muscles, the relationship between CRC and the 30kDa protein remains to be elucidated.(omitted)

• The Korean Journal of Pharmacology
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• v.30 no.1
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• pp.125-136
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• 1994

To characterize the SR Ca-release channel protein complex of crustacean, $^{45}Ca-release,\;[^3H]ryanodine$ binding, and immunoblot studies were carried out in the crayfish and/or lobster skeletal sarcoplasmic reticulum. Bmax and affinity of crayfish SR to ryanodine were lower than those of lobster SR. AMP (5mM) increased $[^3H]ryanodine$ binding significantly in both vesicles (P<0.05). $Mg^{2+}$(5mM) or tetracaine(1mM) inhibited $[^3H]ryanodine$ binding significantly in both vesicles (P<0.001), but ruthenium red $(10\;{\mu}M)$ inhibited it moderately. In SDS polyacrylamide gel electrophoretic analysis of crayfish SR vesicles, there was a high molecular weight band that showed similar mobility with Ca-release channel protein of lobster skeletal SR, but more rapid mobility (HMWBr) than that of rabbit skeletal SR (HMWBS). Immunoblot analysis showed that polyclonal Ab to lobster skeletal SR Ca-release channel protein was react with HMWBr of crayfish skeletal SR, but not with that of HMWBs of rabbit skeletal SR. ^{45}Ca-release from crayfish skeletal SR vesicles was increased by the increase of extravesicular calcium from $1{\mu}M$ to 1mM. This Ca-release phenomenon was similar, but more sensitive in the low concentration of $Ca^{2+}$, compared to that from lobster SR vesicles. AMP (5mM) or caffeine (10mM) did not affect to $^{45}Ca-release.\;^{45}Ca-release$ was inhibited slightly ($3{\sim}8%\;by\;Mg^{2+}$) (5mM) or tetracaine (1mM), and moderately (23%) by high concentration of ruthenium red $(300\;{\mu}M)$. From the above results, it is suggested that SR Ca-release channel protein of crustacean has different properties from that of the rabbit, and similar properties between crayfish and lobster in functional and immunological aspects, but Ca-release via crayfish channel may be more sensitive to calcium.

• The Korean Journal of Physiology and Pharmacology
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• v.9 no.2
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• pp.87-94
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• 2005

It is not clear whether $Ca^{2+}-induced$ $Ca^{2+}$ release from the sarcoplasmic reticulum (SR) is involved in the regulation of atrial natriuretic peptide (ANP) release. Previously, we have shown that nifedipine increased ANP release, indicating that $Ca^{2+}$ entry via voltage-gated L-type $Ca^{2+}$ channel activation decreases ANP release. The purpose of the present study was two-fold: to define the role of SR $Ca^{2+}$ release in the regulation of ANP release and whether $Ca^{2+}$ entry via L-type $Ca^{2+}$ channel is prerequisite for the SR-related effect on ANP release. Experiments were performed in perfused beating rabbit atria. Ryanodine, an inhibitor of SR $Ca^{2+}$ release, increased atrial myocytic ANP release ($8.69{\pm}3.05$, $19.55{\pm}1.09$, $27.31{\pm}3.51$, and $18.91{\pm}4.76$% for 1, 2, 3, and $6{\mu}M$ ryanodine, respectively; all P<0.01) with concomitant decrease in atrial stroke volume and pulse pressure in a dose-dependent manner. In the presence of thapsigargin, an inhibitor of SR $Ca^{2+}$ pump, ryanodine-induced increase in ANP release was not observed. Thapsigargin attenuated ryanodine-induced decrease in atrial dynamic changes. Blockade of L-type $Ca^{2+}$ channel with nifedipine abolished ryanodine-induced increase in ANP release ($0.69{\pm}5.58$% vs. $27.31{\pm}3.51$%; P＜0.001). In the presence of thapsigargin and ryanodine, nifedipine increased ANP release and decreased atrial dynamics. These data suggest that $Ca^{2+}$-induced $Ca^{2+}$ release from the SR is inversely involved in the regulation of atrial myocytic ANP release.

• 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.

• BMB Reports
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• v.29 no.1
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• pp.22-26
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• 1996

Agents that activate or inhibit the $Ca^{2+}$ release channel in cardiac sarcoplasmic reticulum (SR) were tested for their abilities to affect the activity of the SR $Ca^{2+}$-ATPase. Vesicles of junctional SR (heavy SR, HSR) from terminal cisternae were prepared from porcine cardiac muscle by density gradient centrifugation. The steady-state activity of $Ca^{2+}$-ATPases in intact HSR vesicles was/$347{\pm}5\;nmol/min{\cdot}mg$ protein (${\pm}$ SD). When the HSR vesicles were made leaky, the activity was increased to $415{\pm}5\;nmol/min{\cdot}mg$ protein. This increase is probably due to the uncoupling of HSR vesicles. Caffeine (10 mM), an agonist of the SR $Ca^{2+}$ release channel, increased $Ca^{2+}$-ATPase activity in the intact HSR vesicle preparation to $394{\pm}30\;nmol/min{\cdot}mg$ protein. However, caffeine had no significant effect in the leaky vesicle preparation and in the purified $Ca^{2+}$-ATPase preparation. The effect of caffeine on SR $Ca^{2+}$-ATPase was investigated at various concentrations of $Ca^{2+}$. Caffeine increased the pump activity over the whole range of $Ca^{2+}$ concentrations, from $1\;{\mu}M$ to $250\;{\mu}M$, in the intact HSR vesicles. When the SR $Ca^{2+}$-ATPase was inhibited by thapsigargin, no caffeine effect was observed. These results imply that the caffeine effect requires the intact vesicles and that the increase in $Ca^{2+}$-ATPase activity is not due to a direct interaction of caffeine with the enzyme. We propose that the activity of SR $Ca^{2+}$-ATPase is linked indirectly to the activity of the $Ca^{2+}$ release channel (ryanodine receptor) and may depend upon the amount of $Ca^{2+}$ released by the channels.

• The Korean Journal of Physiology and Pharmacology
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• v.6 no.2
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• pp.101-107
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• 2002

In the present study, the postnatal developmental changes in the expressional levels of cardiac sarcoplasmic reticulum (SR) $Ca^{2+}$ regulatory proteins, i.e. $Ca^{2+}-ATPase,$ phospholamban, and $Ca^{2+}$ release channel, were investigated. Both SR $Ca^{2+}-ATPase$ and phospholamban mRNA levels were about 35% of adult levels at birth and gradually increased to adult levels. Protein levels of both SR $Ca^{2+}-ATPase$ and phospholamban, which were measured by quantitative immunoblotting, were closely correlated with the mRNA levels. The initial rates of $Ca^{2+}$ uptake at birth were about 40% of adult rates and also increased gradually during the myocardial development. Consequently, the relative phospholamban/$Ca^{2+}-ATPase$ ratio was 1 in developmental hearts. $Ca^{2+}$ release channel (ryanodine receptor) mRNA was about $50{\sim}60%$ at birth and increased gradually to adult level throughout the postnatal rat heart development. $^3[H]ryanodine$ binding increased gradually during postnatal myocardial development, which was closely correlated with ryanodine mRNA expression levels during the development except the ryanodine mRNA level at birth. These findings indicate that cardiac SR $Ca^{2+}-ATPase,$ phospholamban, and $Ca^{2+}$ release channel are expressed coordinately, which may be necessary for intracellular $Ca^{2+}$ regulation during the rat heart development.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.294-294
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• 1994

To investigate analgesic mechanism of capsaicin and its analogues (capsaicinoids), release of adenosine was measured by high performance liquid chromatography from dorsal spinal cord synaptosomes, Exposure of synaptosomes to K$\^$+/ and morphine produced a dose dependent release of adenosine in the presence of Ca$\^$++/. Capsaicin (0.1, 1, 10 M), and its analogues 6-paradol (1, 10 M), NE-19550 (1, 10, 100 M), DMNE (1, 10, 100 M) and KR 25018 (0.1, 1, 10 M) produced a dose dependent release of adenosine in the presence of Ca$\^$++/. Nifedipine, L-type voltage sensitive calcium channel blocker, inhibited K$\^$+/ (6, 12 mM)- and morphine (10 M)-evoked release of adenosine completely, but inhibited capsaicin, and capsaicinoids-evoked release of adenosine partially. Capsazepine, a novel capsaicin select ive antagonist, blocked only capsaicin and capsaicinoids induced release of adenosine. Therefore, the adenosine release by capsaicin and capsaicinoids having antinociceptive effects involve activation of capsaicin specific receptor and capsaicin sensitive Ca$\^$++/ channel.

• Archives of Pharmacal Research
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• v.25 no.3
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• pp.349-356
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• 2002

The present study was performed to examine the effect of fangchinoline, a bis- benzylisoquinoline alkaloid, which exhibits the characteristics of a $Ca^{2+}$ channel blocker, on cyanide-induced neurotoxicity using cultured rat cerebellar granule neurons. NaCN produced a concentration-dependent reduction of cell viability, which was blocked by MK-801, an N-methyl-D-aspartate (NMDA) receptor antagonist, verapamil, L-type$Ca^{2+}$channel blocker, and L-NAME, a nitric oxide synthase inhibitor. Pretreatment with fangchinoline over a concentration range of 0.1 to 10 $\mu$M significantly decreased the NaCN-induced neuronal cell death, glutamate release into medium, and elevation of $[Ca^{2+}]_i$ and oxidants generation. These results suggest that fangchinoline may mitigate the harmful effects of cyanide-induced neuronal cell death by interfering with $[Ca^{2+}]_i$influx, due to its function as a $Ca^{2+}$ channel blocker, and then by inhibiting glutamate release and oxidants generation.

• BMB Reports
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• v.28 no.2
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• pp.129-137
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• 1995

The ryanodine-receptor $Ca^{2+}$ release channel protein in the sarcoplasmic reticulum membrane of rabbit skeletal muscle plays an important role in muscle exitation-contraction (E-C) coupling. Various types of detergents were tested, including Chaps, cholate, octylglucoside, Zwittergents, Mega-9, Lubrol PX, and Triton X-100 for solubilization of this protein. Among these, Chaps and Triton X-100 were found to optionally solubilize the channel complex. Optimum conditions for this solubilization were pH 7.4 with a salt concentration of 1 M. The addition of phospholipid in the solubilization step helped in stabilizing the protein. The purification of the receptor was performed using sucrose density gradient centrifugation. Various methods [dilution, freeze-thaw, adsorption (Biobeads), and dialysis] were investigated to incorporate the Chaps-solubilized and purified $Ca^{2+}$ release channel protein into liposomes made from different types of phospholipids. Of these, a combined method consisting of a dialysis, freeze-thaw and sonication steps yielded the best results. Reconstituted vesicles produced by this method with 95% phosphatidylcholine (from soybean extract) had good function.