• The Korean Journal of Physiology and Pharmacology
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• v.16 no.1
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• pp.71-77
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• 2012

Mitochondrial dynamics and distribution is critical for their role in bioenergetics and cell survival. We investigated the consequence of altered fission/fusion on mitochondrial function and motility in INS-1E rat clonal ${\beta}$-cells. Adenoviruses were used to induce doxycycline-dependent expression of wild type (WT-Mfn1) or a dominant negative mitofusin 1 mutant (DN-Mfn1). Mitochondrial morphology and motility were analyzed by monitoring mitochondrially-targeted red fluorescent protein. Adenovirus-driven overexpression of WT-Mfn1 elicited severe aggregation of mitochondria, preventing them from reaching peripheral near plasma membrane areas of the cell. Overexpression of DN-Mfn1 resulted in fragmented mitochondria with widespread cytosolic distribution. WT-Mfn1 overexpression impaired mitochondrial function as glucose- and oligomycin-induced mitochondrial hyperpolarization were markedly reduced. Viability of the INS-1E cells, however, was not affected. Mitochondrial motility was significantly reduced in WT-Mfn1 overexpressing cells. Conversely, fragmented mitochondria in DN-Mfn1 overexpressing cells showed more vigorous movement than mitochondria in control cells. Movement of these mitochondria was also less microtubule-dependent. These results suggest that Mfn1-induced hyperfusion leads to mitochondrial dysfunction and hypomotility, which may explain impaired metabolism-secretion coupling in insulin-releasing cells overexpressing Mfn1.

• The Korean Journal of Physiology
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• v.26 no.1
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• pp.55-68
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• 1992

The effects of changes in extracellular $Na^+\;and\;Ca^+$ concentration on the membrane potential and contractility were studied in the antral circular muscle of guinea pig stomach in order to elucidate the existence and the nature of $Na^+/Ca^{2+}$ exchange mechanism. All experiments were performed in tris buffered Tyrode solution which was aerated with 100% $O_2$ and kept at $35^{\circ}C.$ The treatment of $10^{-5}$ ouabain was performed to induce intracellular $Na^+$ loading prior to the start of experiment. The results were as follows: 1. $Na^+$-free Tyrode or high $Ca^{2+}$-Tyrode solution hyperpolarized the membrane potential and induced contracture. The time course of contracture was similar to that of change in membrane potential. 2. The degree of hyperpolarization and the amplitude of contracture decreased in accordance with the increase of extracellular $Na^+$ concentration. 3. $Na^+$-free contracture was developed even after blocking the influence of intrinsic nerves by the pretreatment with atropine, guanethidine and TTX. 4. $Ca^{2+}$-channel blockers(D-600 or $Mn^{2+}$) and the blocker of intracellular $Ca^{2+}$ release from sarcoplasmic reticulum(ryanodine) did not suppress the development of $Na^+$-free contracture. And also, dinitrophenol had no effect on $Na^+$-free contracture. 5. Dose-response relationship between extracellular $Na^+$ concentrations and the magnitude of contractures showed a sigmoid pattern. The slope of straight line from Hill plot was 2.7. 6. In parallel with the increase of extracellular $Ca^{2+}$ concentration, the amplitude of contracture increased dose dependently and was maximum at 8 mM $Ca^{2+}$-Tyrode solution. 7. The relationship between extracellular $Ca^{2+}$ concentrations and the magnitude of contractures showed hyperbolic pattern. The slope of straight line from Hill plot was 1.1. From the above results, it is suggested that $Na^+/Ca^{2+}$ exchange mechanism exists in the antral circular muscle of guinea pig stomach and this mechanism affects the membrane potential electrogenically.

• The Korean Journal of Physiology and Pharmacology
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• v.18 no.5
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• pp.383-390
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• 2014

Gastrointestinal motility consists of phasic slow-wave contractions and the migrating motor complex (MMC). Eupatilin (Stillen$^{(R)}$) has been widely used to treat gastritis and peptic ulcers, and various cytokines and neuropeptides are thought to be involved, which can affect gastrointestinal motility. We performed a study to identify the effects of eupatilin on lower gastrointestinal motility with electromechanical recordings of smooth muscles in the human ileum and colon. Ileum and colon samples were obtained from patients undergoing bowel resection. The tissues were immediately stored in oxygenated Krebs-Ringer's bicarbonate solution, and conventional microelectrode recordings from muscle cells and tension recordings from muscle strips and ileal or colonic segments were performed. Eupatilin was perfused into the tissue chamber, and changes in membrane potentials and contractions were measured. Hyperpolarization of resting membrane potential (RMP) was observed after administration of eupatilin. The amplitude, AUC, and frequency of tension recordings from circular and longitudinal smooth muscle strips and bowel segments of the ileum and colon were significantly decreased after admission of eupatilin. Eupatilin elicited dose-dependent decreases during segmental tension recordings. In conclusion, eupatilin (Stillen$^{(R)}$) showed inhibitory effects on the human ileum and colon. We propose that this drug may be useful for treating diseases that increase bowel motility, but further studies are necessary.

• The Korean Journal of Physiology and Pharmacology
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• v.2 no.1
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• pp.63-68
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• 1998

Nitric oxide (NO)-mediated relaxation in vascular smooth muscle involves not only activation of guanylate cyclase but also hyperpolarization of the membrane. It has been shown that depolarization decreases the [$Ca^{2+}$] sensitivity of myosin light chain kinase in arterial smooth muscle, and nitric oxide (NO)-mediated relaxation was attenuated in this situation. However, why potassium inhibits or attenuates the action of EDRF/NO is not clear. Therefore, we investigated the magnitude of relaxation and cGMP contents using measures known to release NO, such as photorelaxation, photo activated NO-mediated relaxation, and NO-donor (SNP)-mediated relaxation in porcine coronary arterial rings in which contractile conditions were made by different degree of depolarization, i.e., contraction in response to U46619 or U46619 plus KCl. In all cases, the magnitude of relaxation was significantly greater (P＜0.05) in U46619-contracted rings than in U46619+KCl-contracted ones. Although accumulation of cGMP was evident with three measures employed in the present study, no difference was found in cGMP contents between U46619 and U46619+KCl conditions, indicating that the diminished relaxation in KCl containing solution is cGMP-independent mechanism(s). To understand this further, cytosolic $Ca^{2+}$ changes due to NO were compared in rat thoracic aorta by exploiting photoactivated NO using streptozotocin (STZ) that was contracted with either NE or KCl. Fura-3 $[Ca]_{cyt}$ signal caused by NO was small and transient in high $K^+$-, but large and sustained in NE-contracted aorta. The inhibitory potency of STZ expressed in terms of $IC_{50}$ was 5.14 and 3.88 ${\mu}M$ in NE and in high $K^+$, respectively. These results suggest that modification of the cellular mobilization of $Ca^{2+}$ rather than cGMP levels may be an important mechanism for the NO-mediated relaxation when vascular membrane is depolarized, such as atherosclerosis and hypertension.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.1
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• pp.65-74
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• 2017

Here we investigated the central processing mechanisms of mechanical allodynia and found a direct excitatory link with low-threshold input to nociceptive neurons. Experiments were performed on male Sprague-Dawley rats weighing 230-280 g. Subcutaneous injection of interleukin 1 beta ($IL-1{\beta}$) ($1ng/10{\mu}L$) was used to produce mechanical allodynia and thermal hyperalgesia. Intracisternal administration of bicuculline, a gamma aminobutyric acid A ($GABA_A$) receptor antagonist, produced mechanical allodynia in the orofacial area under normal conditions. However, intracisternal administration of bicuculline (50 ng) produced a paradoxical anti-allodynic effect under inflammatory pain conditions. Pretreatment with resiniferatoxin (RTX), which depletes capsaicin receptor protein in primary afferent fibers, did not alter the paradoxical anti-allodynic effects produced by the intracisternal injection of bicuculline. Intracisternal injection of bumetanide, an Na-K-Cl cotransporter (NKCC 1) inhibitor, reversed the $IL-1{\beta}$-induced mechanical allodynia. In the control group, application of GABA ($100{\mu}M$) or muscimol ($3{\mu}M$) led to membrane hyperpolarization in gramicidin perforated current clamp mode. However, in some neurons, application of GABA or muscimol led to membrane depolarization in the $IL-1{\beta}$-treated rats. These results suggest that some large myelinated $A{\beta}$ fibers gain access to the nociceptive system and elicit pain sensation via $GABA_A$ receptors under inflammatory pain conditions.

• The Korean Journal of Physiology and Pharmacology
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• v.4 no.2
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• pp.81-90
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• 2000

The types of $K^+$ channel which determine the pattern of spontaneous action potential (SAP) were investigated using whole-cell variation of patch clamp techniques under current- and voltage-clamp recording conditions in rat clonal pituitary $GH_3$ cells. Heterogeneous pattern of SAP activities was changed into more regular mode with elongation of activity duration and afterhyperpolarization by treatment of TEA (10 mM). Under this condition, exposure of the class III antiarrhythmic agent E-4031 $(5\;{\mu}M)$ to $GH_3$ cells hardly affected SAP activities. On the other hand, the main $GH_3$ stimulator thyrotropin-releasing hormone (TRH) still produced its dual effects (transient hyperpolarization and later increase in SAP frequency) in the presence of TEA. However, addition of $BaCl_2$ (2 mM) in the presence of TEA completely blocked SAP repolarization process and produced membrane depolarization in all tested cells. This effect was observed even in TEA-untreated cells and was not mimicked by higher concentration of TEA (30 mM). Also this barium-induced membrane depolarization effect was still observed after L-type $Ca^{2+}$ channel was blocked by nicardipine $(10\;{\mu}M).$ These results suggest that barium-sensitive current is important in SAP repolarization process and barium itself may have some depolarizing effect in $GH_3$ cells.

• International Journal of Oral Biology
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• v.30 no.3
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• pp.91-98
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• 2005

Gamma-aminobutyric acid (GABA) is known as an inhibitory neurotransmitter in the neurons of the central nervous system. However, its detailed action mechanisms in the rostral gustatory zone of the nucleus tractus solitarius (rNTS) have not been established. The present study was aimed to investigate the distribution, role and action mechanisms of GABA in rNTS. Membrane potentials were recorded by whole cell recordings in isolated brain slices of the rat medulla. Superfusion of GABA resulted in a concentration-dependent reduction in input resistance in the neurons in rNTS. The change in input resistance ws accompanied by response to a depolarizing pulse were diminished by GABA. Superfusion of the slices with either $GABA_A$ agonist, muscimol, $GABA_B$ agonist, baclofen or $GABA_C$ agonist, TACA, decreased input resistance and reduced the nerve activity in association with membrane hyperpolarization. It is suggested that inhibitory signals playa role in sensory processing by the rNTS, in that GABA actions occur through activation of $GABA_A,\;GABA_B\;and\;GABA_C$ receptor. These results suggest that GABA has an inhibitory effect on the rNTS through an activation of $GABA_A,\;GABA_B\;and\;GABA_C$ receptors and that the GABAergic inhibition probably plays an important role in sensory processing by the rNTS.

• Journal of Korean Dental Science
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• v.4 no.2
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• pp.73-78
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• 2011

Purpose: $HCO_3{^-}$ is the most important ion to buffer the acidity of saliva. The transport of $HCO_3{^-}$ is mediated by electrogenic $Na^+/HCO_3{^-}$ cotransporter 1 (NBCe1), which expressed in various tissues including salivary glands, kidney and pancreas, etc. This experiment was performed to investigate regulatory site of NBCe1involved in the pH regulation using various mutants of NBCe1. Materials and Methods: Human parotid gland NBCe1 (hpNBCe1) and mutants by deletion of 1~285 bp and 1~1,035 bp were prepared. After microinjection of each cRNA to oocytes of Xenopus laevis, they were incubated for 2~3 days. The function of each protein was tested by electrophysiological method. Results: When oocytes were exposed to the $HCO_3{^-}$ buffered solution, 1~285 bp deleted mutant hpNBCe1 evoked a marked hyperpolarization ranging from -90 mV to -160 mV (average: -134 mV; n=12) compared to the full length of hpNBCe1. Although 1~1,035 bp deleted mutant hpNBCe1 was also expressed in the plasma membrane, but it did not show any changes of membrane potentials. Conclusion: Our deletion mutant study demonstrated that 1~285 bp of the NBCe1 is the major domain to determine $HCO_3{^-}$ transport ratio.

• The Korean Journal of Physiology and Pharmacology
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• v.6 no.5
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• pp.247-253
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• 2002

Major pelvic ganglia (MPG) neurons are classified into sympathetic and parasympathetic neurons according to the electrophysiological properties; membrane capacitance (Cm), expression of T-type $Ca^{2+}$ channels, and the firing patterns during depolarization. In the present study, function and molecular expression of ATP-sensitive $K^+\;(K_{ATP})$ channels was investigated in MPG neurons of male rats. Only in parasympathetic MPG neurons showing phasic firing patterns, hyperpolarizing changes were elicited by the application of diazoxide, an activator of $K_{ATP}$ channels. Glibenclamide $(10{\mu}M),$ a $K_{ATP}$ channel blocker, completely abolished the diazoxide-induced hyperpolarization. Diazoxide increased inward currents at high $K^+$ (90 mM) external solution, which was also blocked by glibenclamide. The metabolic inhibition by the treatment with mitochondrial respiratory chain inhibitors (rotenone and antimycin) hyperpolarized the resting membrane potential of parasympathetic neurons, which was not observed in sympathetic neurons. The hyperpolarizing response to metabolic inhibition was partially blocked by glibenclamide. RT-PCR analysis revealed that MPG neurons mainly expressed the $K_{ATP}$ channel subunits of Kir6.2 and SUR1. Our results suggest that MPG neurons have $K_{ATP}$ channels, mainly formed by Kir6.2 and SUR1, with phenotype-specificity, and that the conductance through this channel in parasympathetic neurons may contribute to the changes in excitability during hypoxia and/or metabolic inhibition.

• Proceedings of the Korean Biophysical Society Conference
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• 2003.06a
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• pp.39-39
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• 2003

In our previous study, WEHI-231, an immature B cell line, showed intractable increase in [C $a^{2+}$]$_{c}$ after the B-cell receptor (BCR) ligation and treatment with 2-aminoethoxydiphenylborate (2-APB), which was never observed in Bal-17, a mature B cell line (Nam et al., 2003, FEBS Lett). In this study, a whole cell voltage clamp study revealed a specific expression of a novel type of $K^{+}$ current, namely voltage-independent background-type $K^{+}$ channels (IK-bg), in WEHI-231 cells. IK-bg was dramatically increase by the application of 2-APB (50 $\square$M), which induced severe hyperpolarization of WEHI-231 from -45 ㎷ to -90 ㎷, When dialyzed with $Mg^{2+}$ and ATP-free pipette solution, a spontaneous development of IK-bg and membrane hyperpolarization were observed. IK-bg was insensitive to classical $K^{+}$ channel blockers (TEA, glibenclamide, $Ba^{2+}$(1 mM)), whereas blocked by quinine and quinidine in a voltage-dependent manner ($IC_{50}$/=6~9 $\square$M at +60㎷). Phorbol myrstate, a PKC activator, decreased the amplitude of IK-bg. Extracellular acidification (pH 6.5) slightly inhibited IK-bg. Arachidonic acid, riluzole, or hyposmotic stress could not affect the IK-bg after the full development by the intracellular dialysis with Mg-ATP-free solution. In a cell-attached mode of single channel recording from WEHI231, we found two types of voltage-independent $K^{+}$ channels with unitary conductance of 300 pS and 120 pS, respectively. Both channels showed very short mean open times and their open probabilities were increase by the application of 2-APB. In Bal-17 cells, no such $K^{+}$ current was observed in 50 cells tested. In summary, WEHI-231 immature B cells express background $K^{+}$ channels. The pharmacological properties and the large unitary conductance suggest that novel types of two-pore domain $K^{+}$ channels (2-P-K channels) might be expressed in WEHI-231, which may provide an intriguing targets of signal transduction in the immature B lymphocytes.e B lymphocytes.