• Journal of Microbiology and Biotechnology
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• 제15권3호
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• pp.665-671
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• 2005

Initiation and activation of sperm motility are prerequisite processes for the contact and fusion of male and female gametes at fertilization. The phenomena are under the regulation of cAMP and $Ca^{2+}$ in vertebrates and invertebrates. Mammalian sperm requires $Ca^{2+}$ and cAMP for the activation of sperm motility. Cell signaling for the initiation and activation of sperm motility in the ascidians and salmonid fishes has drawn much attention. In the ascidians, the sperm-activating and attracting factors from unfertilized egg require extracellular $Ca^{2+}$ for activating sperm motility and eliciting chemotactic behavior toward the egg. On the other hand, the cAMP-dependent phosphorylation of protein is essential for the initiation of sperm motility in salmonid fishes. A decrease of the environmental $K^+$ concentration surrounding the spawned sperm causes $K^+$ efflux and $Ca^{2+}$ influx through the specific $K^+$ channel and dihydropyridine-sensitive L-/T-type $Ca^{2+}$ channel, respectively, thereby leading to the membrane hyperpolarization. The membrane hyperpolarization induces synthesis of cAMP, which triggers further cell signaling processes, such as cAMP-dependent protein phosphorylation, to initiate sperm motility in salmonid fishes. This article reviews the studies on the physiological mechanisms of sperm motility and its cell signaling in aquatic species.

• The Korean Journal of Physiology and Pharmacology
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• 제6권2호
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• pp.87-91
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• 2002

The aim of this study was to investigate the role of $Ca^{2+}-channel$ blockers in norepinephrine (NE) release from rat hippocampus. Slices and synaptosomes were incubated with $[^3H]-NE$ and the releases of the labelled products were evoked by 25 mM KCl stimulation. Nifedipine, diltiazem, nicardipine, flunarizine and pimozide did not affect the evoked and basal release of NE in the slice. But, diltiazem, nicardipine and flunarizine decreased the evoked NE release with a dose-related manner without any change of the basal release from synaptosomes. Also, a large dose of pimozide produced modest decrement of NE release. ${\omega}-conotoxin$ (CTx) GVIA decreased the evoked NE release in a dose-dependent manner without changing the basal release. And ${\omega}-CTxMVIIC$ decreased the evoked NE release in the synaoptosomes without any effect in the slice, but the effect of decrement was far less than that of ${\omega}-CTxGVIA.$ In interaction experiments with ${\omega}-CTxGVIA,\;{\omega}-CTxMVIIC$ slightly potentiated the effect of ${\omega}-CTxGVIA$ on NE release in the slice and synaptosomal preparations. These results suggest that the NE release in the rat hippocampus is mediated mainly by N-type $Ca^{2+}-channels,$ and that other types such as L-, T- and/or P/Q-type $Ca^{2+}-channels$ could also be participate in this process.

• 고려인삼학회:학술대회논문집
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• 고려인삼학회 2002년도 학술대회지
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• pp.401-415
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• 2002

Lim and his coworkers (1987; 1988; 1989) have also found that all of total Ginseng saponin, panaxadiol-and panaxatriol-type saponins cause the increased secretion of catecholamines (CA) in a $Ca^{2+}$ -dependent fashion from the isolated perfused rabbit adrenal glands through the activation of cholinergic (both nicotinic and muscarinic) receptors. These CA secretory effects are partly due to the direct action on the rabbit adrenomedullary chromaffin cells. However, the present study was designed to examine the effect of total ginseng saponin on CA secretion evoked by activation of cholinergic nicotinic receptors in the isolated perfused model of the rat adrenal gland. Total ginseng saponin given (100 ${\mu}g$/20 min) into an adrenal vein did fail to produce alteration of spontaneous CA release from the rat adrenal medulla. Acetylcholine(5.32 mM)- and DMPP(100 ${\mu}M$, a selective nicotinic receptor agonist)-evoked CA secretory responses were reduced markedly after the pretreatment with the total ginseng saponin at a rate of 100 ${\mu}g$/6.2 ml/20 min, respectively. Pretreatment with total ginseng saponin also depressed greatly high potassium (56 mM, a membrane depolarizing agent)- and Bay-K-8644 (10 ${\mu}M$, a calcium channel activator)-induced CA secretions. Taken together, it is thought that total ginseng saponin can inhibit the releasing effect of CA evoked by nicotinic receptor stimulation from the isolated perfused rat adrenal medulla, which seems to be associated to the direct inhibition of influx through L-type calcium channel into the rat adrenomedullary chromaffin cells. It seems that there is species differences in the adrenomedullary catecholamine secretion between the rabbit and rat.

• The Korean Journal of Physiology and Pharmacology
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• 제2권6호
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• pp.733-742
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• 1998

Background: We have previously reported that not only cGMP but also 8-Br-cGMP or 8-pCPT-cGMP, specific and potent stimulators of cGMP-dependent protein kinase (cGMP-PK), increased basal L-type calcium current $(I_{Ca})$ in rabbit ventricular myocytes. Our findings in rabbit ventricular myocytes were entirely different from the earlier findings in different species, suggesting that the activation of cGMP-PK is involved in the facilitation of $I_{Ca}}$ by cGMP. However, there is no direct evidence that cGMP-PK can stimulate $I_{Ca}}$ in rabbit ventricular myocytes. In this report, we focused on the direct effect of cGMP-PK on $I_{Ca}}$ in rabbit ventricular myocytes. Methods and Results: We isolated single ventricular myocytes of rabbit hearts by using enzymatic dissociation. Regulation of $I_{Ca}}$ by cGMP-PK was investigated in rabbit ventricular myocytes using whole-cell voltage clamp method. $I_{Ca}}$ was elicited by a depolarizing pulse to +10 mV from a holding potential of -40 mV. Extracellular 8-(4-Chlorophenylthio)-guanosine-3',5'-cyclic monophosphate (8-pCPT-cGMP), potent stimulator of cGMP-dependent protein kinase (cGMP-PK), increased basal $I_{Ca}}$. cGMP-PK also increased basal $I_{Ca}}$. The stimulation of basal $I_{Ca}}$ by cGMP-PK required both 8-Br-cGMP in low concentration and intracellular ATP to be present. The stimulation of basal $I_{Ca}}$ by cGMP-PK was blocked by heat inactivation of the cGMP-PK and by bath application of 8-(4-chlorophenylthio)-guanosine-3',5'-cyclic monophosphate, Rp-isomer (Rp-pCPT-cGMP), a phosphodiesterase-resistant cGMP-PK inhibitor. When $I_{Ca}}$ was increased by internal application of cGMP-PK, IBMX resulted in an additional stimulation of $I_{Ca}}$. In the presence of cGMP-PK, already increased $I_{Ca}}$ was potentiated by bath application of isoprenaline or forskolin or intracellular application of cAMP. Conclusions: We present evidence that cGMP-PK stimulated basal $I_{Ca}}$ by a direct phosphorylation of L-type calcium channel or associated regulatory protein in rabbit ventricular myocytes.

• 한국생물물리학회:학술대회논문집
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• 한국생물물리학회 2003년도 정기총회 및 학술발표회
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• pp.19-19
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• 2003

We have developed a cardiac cell model (Kyoto Model) for the sinoatrial node and ventricle, which is composed of a common set of kinetic equations of membrane ionic currents, Ca$\^$2+/dynamics of sarcoplasmic reticulum and contractile protein. To expand this model by including metabolic pathways, the intracellular ATP metabolism, which is pivotal in cardiac excitation - contraction coupling, was incorporated. ATP consumption by the sarcolemmal Na$\^$+/ pump and the Ca pump in the sarcoplasmic reticulum were calculated with stoichiometry of 3Na:2K:1ATP and 2Ca:1ATP, respectively. ATP consumption by contraction was estimated according to experimental data. Dependence of contraction on ATP and inorganic phosphate was modeled, based on data of skinned cardiac fiber. in production by mitochondrial oxidative phosphorylation was modified from Korzeniewski '||'&'||' Zoladz (2001), and creatine kinase and adenylate kinase reactions were incorporated. ATP dependence of ATP-sensitive K channel and L type Ca channel were also included.

• The Korean Journal of Physiology and Pharmacology
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• 제18권5호
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• pp.431-439
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• 2014

The aim of the present study was to investigate whether ginsenoside-Rb2 (Rb2) can affect the secretion of catecholamines (CA) in the perfused model of the rat adrenal medulla. Rb2 ($3{\sim}30{\mu}M$), perfused into an adrenal vein for 90 min, inhibited ACh (5.32 mM)-evoked CA secretory response in a dose- and time-dependent fashion. Rb2 ($10{\mu}M$) also time-dependently inhibited the CA secretion evoked by DMPP ($100{\mu}M$, a selective neuronal nicotinic receptor agonist) and high $K^+$ (56 mM, a direct membrane depolarizer). Rb2 itself did not affect basal CA secretion (data not shown). Also, in the presence of Rb2 ($50{\mu}g/mL$), the secretory responses of CA evoked by veratridine (a selective $Na^+$ channel activator ($50{\mu}M$), Bay-K-8644 (an L-type dihydropyridine $Ca^{2+}$ channel activator, $10{\mu}M$), and cyclopiazonic acid (a cytoplasmic $Ca^{2+}$-ATPase inhibitor, $10{\mu}M$) were significantly reduced, respectively. Interestingly, in the simultaneous presence of Rb2 ($10{\mu}M$) and L-NAME (an inhibitor of NO synthase, $30{\mu}M$), the inhibitory responses of Rb2 on ACh-evoked CA secretory response was considerably recovered to the extent of the corresponding control secretion compared with the inhibitory effect of Rb2-treatment alone. Practically, the level of NO released from adrenal medulla after the treatment of Rb2 ($10{\mu}M$) was greatly elevated compared to the corresponding basal released level. Collectively, these results demonstrate that Rb2 inhibits the CA secretory responses evoked by nicotinic stimulation as well as by direct membrane-depolarization from the isolated perfused rat adrenal medulla. It seems that this inhibitory effect of Rb2 is mediated by inhibiting both the influx of $Ca^{2+}$ and $Na^+$ into the adrenomedullary chromaffin cells and also by suppressing the release of $Ca^{2+}$ from the cytoplasmic calcium store, at least partly through the increased NO production due to the activation of nitric oxide synthase, which is relevant to neuronal nicotinic receptor blockade.

• Molecules and Cells
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• 제25권2호
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• pp.242-246
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• 2008

To assess the role of $\alpha_{1G}$ T-type $Ca^{2+}$ channels in neuropathic pain after L5 spinal nerve ligation, we examined behavioral pain susceptibility in mice lacking $Ca_{V}3.1$ (${\alpha}_{1G}{^{-/-}}$), the gene encoding the pore-forming units of these channels. Reduced spontaneous pain responses and an increased threshold for paw withdrawal in response to mechanical stimulation were observed in these mice. The ${{\alpha}_{1G}}^{-/-}$ mice also showed attenuated thermal hyperalgesia in response to both low-(IR30) and high-intensity (IR60) infrared stimulation. Our results reveal the importance of ${\alpha}_{1G}$ T-type $Ca^{2+}$ channels in the development of neuropathic pain, and suggest that selective modulation of ${\alpha}_{1G}$ subtype channels may provide a novel approach to the treatment of allodynia and hyperalgesia.

• Molecules and Cells
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• 제21권1호
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• pp.52-62
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• 2006

In previous reports we demonstrated that ginsenosides, active ingredients of Panax ginseng, affect some subsets of voltage-dependent $Ca^{2+}$ channels in neuronal cells expressed in Xenopus laevis oocytes. However, the major component(s) of ginseng that affect cloned $Ca^{2+}$ channel subtypes such as ${\alpha}_{1C}$(L)-, ${\alpha}_{1B}$(N)-, ${\alpha}_{1A}$(P/Q)-, ${\alpha}_{1E}$(R)- and ${\alpha}_{1G}$(T) have not been identified. Here, we used the two-microelectrode voltage clamp technique to characterize the effects of ginsenosides and ginsenoside metabolites on $Ba^{2+}$ currents ($I_{Ba}$) in Xenopus oocytes expressing five different $Ca^{2+}$ channel subtypes. Exposure to ginseng total saponins (GTS) induced voltage-dependent, dose-dependent and reversible inhibition of the five channel subtypes, with particularly strong inhibition of the ${\alpha}_{1G}$-type. Of the various ginsenosides, $Rb_1$, Rc, Re, Rf, $Rg_1$, $Rg_3$, and $Rh_2$, ginsenoside $Rg_3$ also inhibited all five channel subtypes and ginsenoside $Rh_2$ had most effect on the ${\alpha}_{1C}$- and ${\alpha}_{1E}$-type $Ca^{2+}$ channels. Compound K (CK), a protopanaxadiol ginsenoside metabolite, strongly inhibited only the ${\alpha}_{1G}$-type of $Ca^{2+}$ channel, whereas M4, a protopanaxatriol ginsenoside metabolite, had almost no effect on any of the channels. $Rg_3$, $Rh_2$, and CK shifted the steady-state activation curves but not the inactivation curves in the depolarizing direction in the ${\alpha}_{1B}$- and ${\alpha}_{1A}$-types. These results reveal that $Rg_3$, $Rh_2$ and CK are the major inhibitors of $Ca^{2+}$ channels in Panax ginseng, and that they show some $Ca^{2+}$ channel selectivity.

• 한국의학물리학회지:의학물리
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• 제17권2호
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• pp.96-104
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• 2006

본 연구자를 위시한 많은 연구자에 의해 칼슘이 N형 칼슘통로의 비활성화를 촉진시킨다는 것이 보고되었다. 그러나 칼슘에 의한 비활성화 촉진 효과가 고전적인 칼슘의존성 기전에 의해 기인하는지는 아직 확실하지 않다. L형 칼슘통로의 칼슘의존성 비활성화기전을 밝히기 위하여 지금까지 사용해온 방법의 하나는 세포내, 외의 칼슘농도를 변화시켜보는 것이다. 그러므로 본 연구에서는 칼슘의존성 비활성화기전의 존재 여부를 알아보기 위하여 2가 양이온을 1가 양이온인 메틸아민($MA^+$)으로 치환하였다. 선행 연구를 통해 우리는 5초 동안의 긴 저분극 자극 시 바륨과 칼슘을 사용하여 얻은 전류에서 모두 빠른 성분(${\tau}{\sim}150ms$)과 느린 성분(${\tau}{\sim}2,500ms$)의 비활성화가 있음을 알 수 있었다. 본 연구에서 세포외 2가 양이온의 농도가 0이 되도록 하였을 때 빠른 비활성화가 소실된 반면 느린 비활성화에는 영향이 거의 없었다. 또한 바륨를 사용하였을 때보다 10 mV씩 과분극시킨 전압에서의 메틸암모늄 전류 데이터를 비교하여 보았을 때 느린 비활성화의 시정수가 서로 잘 일치하였으며 이 시정수는 막전압이 저분극될수록 감소하는 막전압의존성 비활성화의 특성을 보였다. 본 연구결과와 선행연구의 결과를 종합하여 볼 때 세포외 2가 양이온의 존재는 N형 칼슘통로의 빠른 비활성화가 일어나기 위하여 필수적인 조건이며 이러한 2가 양이온의존성 비활성화기전은 기존의 칼슘의존성 또는 막전압의존성 기전과 다르다는 가설을 제안한다.

• The Korean Journal of Physiology and Pharmacology
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• 제23권6호
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• pp.539-547
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• 2019

Anoctamin 5 (ANO5)/TMEM16E belongs to a member of the ANO/TMEM16 family member of anion channels. However, it is a matter of debate whether ANO5 functions as a genuine plasma membrane chloride channel. It has been recognized that mutations in the ANO5 gene cause many skeletal muscle diseases such as limb girdle muscular dystrophy type 2L (LGMD2L) and Miyoshi muscular dystrophy type 3 (MMD3) in human. However, the molecular mechanisms of the skeletal myopathies caused by ANO5 defects are poorly understood. To understand the role of ANO5 in skeletal muscle development and function, we silenced the ANO5 gene in C2C12 myoblasts and evaluated whether it impairs myogenesis and myotube function. ANO5 knockdown (ANO5-KD) by shRNA resulted in clustered or aggregated nuclei at the body of myotubes without affecting differentiation or myotube formation. Nuclear positioning defect of ANO5-KD myotubes was accompanied with reduced expression of Kif5b protein, a kinesin-related motor protein that controls nuclear transport during myogenesis. ANO5-KD impaired depolarization-induced $[Ca2^{+}]_i$ transient and reduced sarcoplasmic reticulum (SR) $Ca^{2+}$ storage. ANO5-KD resulted in reduced protein expression of the dihydropyridine receptor (DHPR) and SR $Ca^{2+}-ATPase$ subtype 1. In addition, ANO5-KD compromised co-localization between DHPR and ryanodine receptor subtype 1. It is concluded that ANO5-KD causes nuclear positioning defect by reduction of Kif5b expression, and compromises $Ca^{2+}$ signaling by downregulating the expression of DHPR and SERCA proteins.