• Title/Summary/Keyword: Whole-cell patch-clamp technique

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Inactivation of N-Type Calcium Current in Rat Sympathetic Neurons

  • Lee, Mi-Sun;Goo, Yong-Sook
    • Proceedings of the Korean Biophysical Society Conference
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    • 2001.06a
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    • pp.26-26
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    • 2001
  • Inactivation of N-type calcium current has been reported to be both voltage dependent and Ca$\^$2+/ dependent. We have investigated the effects of Ba$\^$2+/ and Ca$\^$2+/ on N-channel inactivation in rat superior cervical ganglion neurons using the whole cell configuration of patch clamp technique. Inactivation was larger in Ca$\^$2+/ than in Ba$\^$2+/ even with 20 mM BAPTA.(omitted)

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Changes in Intracellular $Ca^{2+}$ Concentration Induced by L-Type $Ca^{2+}$ Channel Current in Guinea-Pig Gastric Myocytes

  • Kim, Ki-Whan
    • Proceedings of the Korean Biophysical Society Conference
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    • 1997.07a
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    • pp.17-17
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    • 1997
  • We investigated the relationship between the voltage-operated calcium channel current and the corresponding [Ca$^{2+}$]i change (Ca$^{2+}$-transient) in guinea-pig gastric myocyte. Fluorescence microspectroscopy was combined with conventional whole-cell patch clamp technique and fura-2 (80 $\mu$M) was added into the CsCl-rich pipette solution.(omitted)

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The Effect of Papaverine on the Calcium-dependent $K^+$ Current in Rat Basilar Smooth Muscle Cells

  • Bai, Guang-Yi;Cho, Jae-Woo;Han, Dong-Han;Yang, Tae-Ki;Gwak, Yong-Geun;Kim, Chul-Jin
    • Journal of Korean Neurosurgical Society
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    • v.38 no.5
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    • pp.375-379
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    • 2005
  • Objective : Papaverine has been used in treating vasospasm following subarachnoid hemorrhage[SAH]. However, its action mechanism for cerebral vascular relaxation is not clear. Potassium channels are closely related to the contraction and relaxation of cerebral smooth muscle. Therefore, to identify the role of potassium and calcium channels in papaverine-induced vascular relaxation, we examine the effect of papaverine on potassium channels in freshly isolated smooth muscle cells from rat basilar artery. Methods : The isolation of rat basilar smooth muscle cells was performed by special techniques. The whole cell currents were recorded by whole cell patch clamp technique in freshly isolated smooth muscle cells from rat basilar artery. Papaverine was added to the bath solution. Results : Papaverine of $100{\mu}M$ into bath solution increased the amplitude of the outward $K^+$ current which was completely blocked by BKCa[large conductance calcium dependent potassium channels]blocker, IBX[iberiotoxin], and calcium chealator, BAPTA[l,2-bis[o-aminophenoxy]ethane-N,N,N',N'-tetraacetic acid], in whole cell mode. Conclusion : These results strongly suggest that potassium channels may play roles in papaverine-induced vascular relaxation in rat basilar artery.

[$Ca^{2+}-activated\;Cl^-$ Current in Gastric Antral Myocytes

  • Lee, Moo-Yeol;Bang, Hyo-Weon;Uhm, Dae-Yong;Rhee, Sang-Don
    • The Korean Journal of Physiology
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    • v.28 no.2
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    • pp.143-150
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    • 1994
  • The whole-cell mode of the patch clamp technique was used to study $Ca^{2+}-activated\;Cl^-\;current$ $(I_{Cl_{Ca}})$ in gastric antral myocytes. Extracellular application of caffeine evoked $Ca^{2+}-activated\;current$. In order to isolate the chloride current from background current, all known systems were blocked with specific blockers. The current-voltage relationship of caffeine-induced current showed outward rectification and it reversed at around $E_{Cl^-}$. The shift of reversal potential upon the alteration of external and internal chloride concentrations was well fitted with results which were calculated by the Nernst equation. Extracellular addition of N-phenylanthranilic acid and niflumic acid which are known anion channel blockers abolished the caffeine induced current. Intracellular application of a high concentration of EGTA also abolished this current. Application of c-AMP, c-GMP, heparin, or $AIF^-_4$ made no remarkable changes to this current. Sodium replacement with the impermeable cation N-methylglucamine or with $Cd^{2+}$ rarely affected this current. From the above results it is suggested that the caffeine induced current was a $Cl^-$ current and it was activated by intracellular $Ca^{2+}$.

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Effects of Prostaglandin $F_{2{\alpha}}$ on Membrane Potentials and $K^+$ Currents in Rabbit Middle Cerebral Arterial Cells

  • Kim, Na-Ri;Han, Jin;Kim, Won-Gue;Kim, Eui-Yong
    • The Korean Journal of Physiology and Pharmacology
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    • v.4 no.4
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    • pp.301-309
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    • 2000
  • The purpose of our investigation was to examine the effects of prostaglandin $F_{2{\alpha}}\;(PGF_{2{\alpha}})$ on membrane potentials, $Ca^{2+}-activated\;K^+\;(K_{Ca})$ channels, and delayed rectifier $K^+(K_V)$ channels using the patch-clamp technique in single rabbit middle cerebral arterial smooth muscle cells. $PGF_{2{\alpha}}$ significantly hyperpolarized membrane potentials and increased outward whole-cell K currents. $PGF_{2{\alpha}}$ increased open-state probability of $K_{Ca}$ channels without the change of the open and closed kinetics. $PGF_{2{\alpha}}$ increased the amplitudes of $K_V$ currents with a leftward shift of activation and inactivation curves and a decrease of activation time constant. Our results suggest that the activation of $K_{Ca}$ and $K_V$ channels, at least in part, may lead to attenuate or counteract vasoconstriction by $PGF_{2{\alpha}}$ in middle cerebral artery.

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Effects of Phenylephrine on the Excitability of Medial Vestibular Nuclear Neurons in Rats

  • Jeong, Han-Seong;Huh, Hae-Ryong;Jang, Myung-Joo;Hong, Seol-Hee;Jang, Su-Jeong;Park, Jin;Lee, Seung-Han;Kim, Jae-Ha;Park, Jong-Seong
    • The Korean Journal of Physiology and Pharmacology
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    • v.10 no.3
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    • pp.131-135
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    • 2006
  • Coeruleo-vestibular pathway which connects locus coeruleus and vestibular nuclei is noradrenergic. This study was designed to elucidate the effects of phenylephrine on the spontaneous activity of acutely isolated medial vestibular nuclear neurons of rat by whole-cell patch-clamp technique. Sprague-Dawley rats, aged 14 to 16 days, were used. After enzymatic digestion, dissociated medial vestibular neurons were transferred to a recording chamber mounted on an inverted microscope, and spontaneous action potentials were recorded by standard patch-clamp techniques. In current-clamp mode, the frequency of spontaneous action potential of medial vestibular nuclear neurons was decreased by phenylephrine (n=15). Phenylephrine increased the amplitude of afterhyperpolarization without changes in the resting membrane potential and spike width. In voltage-clamp mode, the whole potassium currents of the medial vestibular nuclear neurons were increased by phenylephrine (n=12). These experimental results suggest that ${\alpha}-receptor$ mediates the inhibitory effects on the neuronal activity of the medial vestibular nuclear neuron.

Effects of Noradrenaline on the Spontaneous Contraction and Ionic Current in the Antral Circular Muscle of Guinea-pig Stomach

  • Jun, Jae-Yeoul;Lee, Sang-Jin;Kim, Sung-Joon;Suh, Jae-Yul;So, In-Suk;Hwang, Sang-Ik;Kim, Ki-Whan
    • The Korean Journal of Physiology
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    • v.27 no.2
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    • pp.115-122
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    • 1993
  • There is evidence that noradrenaline enhances spontaneous contractions dose-dependently in guinea-pig antral circular muscle. To investigate the mechanism of this excitatory action, slow waves and membrane currents were recorded using conventional microelectrode techniques in muscle strips and the whole cell patch clamp technique in isolated gastric myocytes. On recording slow waves, noradrenaline $(10^{-5}\;M)$ induced the hyperpolarization of the membrane potential, although the shape of the slow waves became tall and steep. Also, spike potentiaIs occurred at the peaks of slow waves. These changes were completely reversed by administration of phentolamine $(10^{-5}\;M),\;an\;{\alpha}-adrenoceptor$ blocker. Noradrenaline-induced hyperpolarization was blocked by apamin $(10^{-7}\;M)$, a blocker of a class of $Ca^{2+}\;-dependent\;K^+$ channels. To investigate the mechanisms for these effects, we performed whole cell patch clamp experiments. Norndrenaline increased voltage-dependent $Ca^{2+}$ currents in the whole range of test potentials. Noradrenaline also increased $Ca^{2+}\;-dependent\;K^+$\;currents, and this effects was abolished by apamin. These results suggest that the increase in amplitude and the generation of spike potentials on slow waves was caused by the activation of voltage-dependent $Ca^{2+}$ channel via adrenoceptors, and hyperpolarization of the membrane potential was mediated by activation of apamin-sensitive $Ca^{2+}\;-dependent\;K^+\;channels$.

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Physiological and Pharmacological Characterization of Glutamate and GABA Receptors in the Retina

  • Yang, Xiong-Li;Shen, Ying;Han, Ming-Hu;Lu, Tao
    • The Korean Journal of Physiology and Pharmacology
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    • v.3 no.5
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    • pp.461-469
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    • 1999
  • Glutamate and ${\gamma}-aminobutyric$ acid (GABA) are major excitatory and inhibitory neurotransmitters in the vertebrate retina, respectively. Using the whole-cell patch clamp technique and a rapid solution changer, glutamate and GABA receptors have been extensively investigated in carp retina. Glutamate receptors on both horizontal and amacrine cells may be an AMPA preferring subtype, which predominantly consists of flop splice variants. $GABA_A$ and $GABA_C$ receptors coexist in bipolar cells and they both show significant desensitization. Kinetics analysis demonstrated that activation, deactivation and desensitization of the $GABA_C$ receptor-mediated response of these cells are overall slower than those of the $GABA_A$ response. Endogenous modulator $Zn^{2+}$ in the retina was found to differentially modulate the kinetic characteristics of the $GABA_C$ and $GABA_A$ responses.

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Effect of Capsaicin on Delayed Rectifier $K^+$ Current in Adult Rat Dorsal Root Ganglion Neurons

  • Hahn, Jung-Hyun;Chung, Sung-Kwon;Bang, Hyo-Weon
    • The Korean Journal of Physiology and Pharmacology
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    • v.4 no.1
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    • pp.9-14
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    • 2000
  • $K^+$ currents play multiple roles in the excitability of dorsal root ganglion (DRG) neurons. Influences on these currents change the shape of the action potential, its firing threshold and the resting membrane potential. In this study, whole cell configuration of patch clamp technique had been applied to record the blocking effect of capsaicin, a lipophilic alkaloid, on the delayed rectifier $K^+$ current in cultured small diameter DRG neurons of adult rat. Capsaicin reduced the amplitude of $K^+$ current in dose dependent manner, and the concentration-dependence curve was well described by the Hill equation with $K_D$ value of $19.1{\mu}M.$ The blocking effect of capsaicin was reversible. Capsaicin $(10 {\mu}M)$ shifted the steady-state inactivation curve in the hyperpolarizing direction by about 15 mV and increased the rate of inactivation. The voltage dependence of activation was not affected by capsaicin. These multiple effects of capsaicin may suggest that capsaicin bind to the region of $K^+$ channel, participating in inactivation process.

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Effect of Cyclic GMP on the Calcium Current in Rabbit Ventricular Myocytes

  • Han, Jin;Leem, Chae-Hun;Ahn, Chul-Soo;So, In-Suk;Kim, Eui-Yong;Ho, Won-Kyung;Earm, Yung-E
    • The Korean Journal of Physiology
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    • v.27 no.2
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    • pp.151-162
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    • 1993
  • In order to investigate the effect of intracellular cyclic GMP on calcium current the whole-cell patch clamp technique with internal perfusion method was used in isolated ventricular myocytes of the rabbit. Cyclic GMP, 8-bromo-cyclic GMP, cyclic AMP, isoprenaline and forskolin were perfused into cells and their effects on calcium current were analysed by applying depolarizing step pulses of + 10 mV in amplitude far 300 msec from holding potential of - 40 mV. Not only cyclic AMP $(100\;{\mu}M)$ but also cyclic GMF $(100\;{\mu}M)$ increased the basal calcium current. 8-Bromo-cyclic GMP $(100\;{\mu}M)$, a good stimulator of the cyclic GMP-dependent protein kinase, also increased the basal calcium current and its peak amplitude of calcium current was larger than that in the presence of cyclic AMP or cyclic GMP alone. In the presence of $100\;{\mu}M$ cyclic GMP or $100\;{\mu}M$ 8-bromo-cyclic GMP, already augmented calcium current was potentiated by intracellular application of $100\;{\mu}M$ cyclic AMP or $1\;{\mu}M$ isoprenaline or $1\;{\mu}M$ forskolin. In the presence of cyclic GMP, acetylcholine reduced the calcium current only when the calcium current was increased by isoprenaline. From the above results it could be concluded that intracellular perfusion with cyclic GMP increases the basal calcium current via a mechanism involving a cyclic GMP-dependent protein kinase.

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