• The Korean Journal of Physiology
• /
• 제27권2호
• /
• pp.115-122
• /
• 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 $Ca^{2+}\;-dependent\;K^+\;channels$.

• Journal of Ginseng Research
• /
• 제35권2호
• /
• pp.219-225
• /
• 2011

Korean red ginseng (KRG) is a valuable and important traditional medicine in East Asian countries and is currently used extensively for botanical products in the world. KRG has both stimulatory and inhibitory effects on the central nervous system (CNS) suggesting its complicated action mechanisms. The substantia gelatinosa (SG) neurons of the trigeminal subnucleus caudalis (Vc) are involved in orofacial nociceptive processing. Some studies reported that KRG has antinociceptive effects, but there are few reports of the functional studies of KRG on the SG neurons of the Vc. In this study, a whole cell patch clamp study was performed to examine the action mechanism of a KRG extract on the SG neurons of the Vc from juvenile mice. KRG induced short-lived and repeatable inward currents on all the SG neurons tested in the high chloride pipette solution. The KRG-induced inward currents were concentration dependent and were maintained in the presence of tetrodotoxin, a voltage gated $Na^+$ channel blocker. The KRG-induced inward currents were suppressed by 6-cyano-7-nitroquinoxaline-2,3-dione, a non-N-methyl-D-aspartate (NMDA) glutamate receptor antagonist and/or picrotoxin, a gamma-aminobutyric acid $(GABA)_A$ receptor antagonist. However, the inward currents were not suppressed by d,l-2-amino-5-phosphonopentanoic acid, an NMDA receptor antagonist. These results show that KRG has excitatory effects on the SG neurons of the Vc via the activation of non-NMDA glutamate receptor as well as an inhibitory effect by activation of the $GABA_A$ receptor, indicating the KRG has both stimulatory and inhibitory effects on the CNS. In addition, KRG may be a potential target for modulating orofacial pain processing.

• Experimental and Molecular Medicine
• /
• 제50권12호
• /
• pp.11.1-11.9
• /
• 2018

Estrogen has diverse effects on cardiovascular function, including regulation of the contractile response to vasoactive substances such as serotonin. The serotonin system recently emerged as an important player in the regulation of vascular tone in humans. However, hyperreactivity to serotonin appears to be a critical factor for the pathophysiology of hypertension. In this study, we examined the modulatory mechanisms of estrogen in serotonin-induced vasoconstriction by using a combinatory approach of isometric tension measurements, molecular biology, and patch-clamp techniques. $17{\beta}$-Estradiol (E2) elicited a significant and concentration-dependent relaxation of serotonin-induced contraction in deendothelialized aortic strips isolated from male rats. E2 triggered a relaxation of serotonin-induced contraction even in the presence of tamoxifen, an estrogen receptor antagonist, suggesting that E2-induced changes are not mediated by estrogen receptor. Patch-clamp studies in rat arterial myocytes showed that E2 prevented Kv channel inhibition induced by serotonin. Serotonin increased Src activation in arterial smooth muscle required for contraction, which was significantly inhibited by E2. The estrogen receptor-independent inhibition of Src by E2 was confirmed in HEK293T cells that do not express estrogen receptor. Taken together, these results suggest that estrogen exerts vasodilatory effects on serotonin-precontracted arteries via Src, implying a critical role for estrogen in the prevention of vascular hyperreactivity to serotonin.

• 대한의생명과학회지
• /
• 제15권2호
• /
• pp.127-133
• /
• 2009

This study was designed to investigate the effects of high concentration of (-)-epigallocatechin-3-gallate(EGCG) on the neuronal activity of rat substantia nigra dopaminergic neurons. Sprague-Dawley rats aged 14 to 16 days were decapitated under ether anesthesia. After treatment with pronase and thermolysin, the dissociated dopaminergic neurons were transferred into a chamber on an inverted microscope. Spontaneous action potentials and potassium currents were recorded by standard patch-clamp techniques under current and voltage-clamp modes respectively. 18 dopaminergic neurons(80%) revealed inhibitory responses to 40 and 100 ${\mu}M$ of EGCG and 4 neurons(20%) did not respond to EGCG. The spike frequency and resting membrane potential of these cells were decreased by EGCG. The amplitude of afterhyperpolarization was increased by EGCG. Whole potassium currents of dopaminergic neurons were increased by EGCG(n=10). These experimental results suggest that high concentration EGCG decreases the neuronal activity of the dopaminergic neurons by altering the resting membrane potential and afterhyperpolarization.

• The Korean Journal of Physiology and Pharmacology
• /
• 제9권4호
• /
• pp.209-213
• /
• 2005

Interstitial cells of Cajal (ICCs) are the pacemaker cells that generate slow waves in the gastrointestinal (GI) tract. In the present study, we investigated the effect of mitochondrial ATP-sensitive potassium (mitoKATP) channel on pacemaking activity in cultured ICCs from murine small intestine by using whole-cell patch clamp techniques. Under current clamp mode, at 10μM glibenclamide, there was no change in pacemaking activity of ICCs. At $30{\mu}M$ glibenclamide, an inhibitor of the ATP sensitive $K^+$ channels, we could find two examples. If pacemaking activity of ICCs was irregulating, pacemaking activity of ICCs was changed into regulating and if in normal conditions, membrane potential amplitude was increased. At $50{\mu}M$ glibenclamide, the resting membrane potential was depolarized. At 3mM 5-HDA, an inhibitor of the mitoKATP channels, inhibited the pacemaking activity of ICCs. Both the amplitude and the frequency were decreased. At 5 mM 5-HDA, both the amplitude and the frequency were completely abolished. Diazoxide, an opener of the mitoKATP channels, was applied to examine its effect on pacemaking activity of ICCs. At $50{\mu}M$ concentration, the pacemaking activity of ICCs was inhibited. Both the amplitude and the frequency were decreased. At 1 mM concentration, both the amplitude and the frequency were completely abolished and the resting membrane potential was shaked.These results indicate that mitoKATP channel has an important role in pacemaking activity of ICCs.

• 대한의생명과학회지
• /
• 제16권1호
• /
• pp.46-52
• /
• 2010

This study was designed to investigate the effects of sphingosine-1-phosphate on the neuronal activity of rat medial vestibular nuclear neurons. Sprague-Dawley rats aged 14 to 16 days were decapitated under ether anesthesia. After treatment with pronase and thermolysin, the dissociated medial vestibular nuclear neurons were transferred into a chamber on an inverted microscope. Spontaneous action potentials and potassium currents were recorded by standard patch-clamp techniques under current and voltage-clamp modes respectively. 15 medial vestibular nuclear neurons revealed excitatory responses to 1 and $5\;{\mu}M$ of sphingosine-1-phosphate. The spike frequency and resting membrane potential of these cells were increased by sphingosine-1-phosphate. The amplitude of afterhyperpolarization was decreased by sphingosine-1-phosphate. Whole potassium currents of medial vestibular nuclear neurons were decreased by sphingosine-1-phosphate (n=12). Sphingosine-1-phosphate did not affect the charybdotoxin-treated potassium currents. These experimental results suggest that sphingosine-1-phosphate increases the neuronal activity of the medial vestibular nuclear neurons by altering the resting membrane potential and afterhyperpolarization.

• Journal of Ginseng Research
• /
• 제39권2호
• /
• pp.169-177
• /
• 2015

Background: Ginsenoside Rd (GSRd), one of the most abundant ingredients of Panax ginseng, protects the heart via multiple mechanisms including the inhibition of $Ca^{2+}$ influx.We intended to explore the effects of GSRd on L-type $Ca^{2+}$ current ($I_{Ca,L}$) and define the mechanism of the suppression of $I_{Ca,L}$ by GSRd. Methods: Perforated-patch recording and whole-cell voltage clamp techniques were applied in isolated rat ventricular myocytes. Results: (1) GSRd reduced $I_{Ca,L}$ peak amplitude in a concentration-dependent manner [half-maximal inhibitory concentration $(IC_{50})=32.4{\pm}7.1{\mu}mol/L$] and up-shifted the current-voltage (I-V) curve. (2) GSRd ($30{\mu}mol/L$) significantly changed the steady-state activation curve of $I_{Ca,L}$ ($V_{0.5}:-19.12{\pm}0.68$ vs. $-6.26{\pm}0.38mV$; n = 5, p < 0.05) and slowed down the recovery of $I_{Ca,L}$ from inactivation [the time content (${\zeta}$) from 91 ms to 136 ms, n = 5, p < 0.01]. (3) A more significant inhibitive effect of GSRd ($100{\mu}mol/L$) was identified in perforated-patch recording when compared with whole-cell recording [$65.7{\pm}3.2%$ (n = 10) vs. $31.4{\pm}5.2%$ (n = 5), p < 0.01]. (4) Pertussis toxin ($G_i$ protein inhibitor) completely abolished the $I_{Ca,L}$ inhibition induced by GSRd. There was a significant difference in inhibition potency between the two cyclic adenosine monophosphate elevating agents (isoprenaline and forskolin) prestimulation [$55{\pm}7.8%$ (n = 5) vs. $17.2{\pm}3.5%$ (n = 5), p < 0.01]. (5) 1H-[1,2,4]Oxadiazolo[4,3-a]-quinoxalin-1-one (a guanylate cyclase inhibitor) and N-acetyl-$\small{L}$-cysteine (a nitric oxide scavenger) partly recovered the $I_{Ca,L}$ inhibition induced by GSRd. (6) Phorbol-12-myristate-13-acetate (a protein kinase C activator) and GF109203X (a protein kinase C inhibitor) did not contribute to the inhibition of GSRd. Conclusion: These findings suggest that GSRd could inhibit $I_{Ca,L}$ through pertussis toxin-sensitive G protein ($G_i$) and a nitric oxide-cyclic guanosine monophosphate-dependent mechanism.

• The Korean Journal of Physiology and Pharmacology
• /
• 제9권4호
• /
• pp.203-207
• /
• 2005

Electrical rhythmicity in the gastrointestinal (GI) muscles is generated by pacemaker cells, known as interstitial cells of Cajal (ICC). In the present study, we investigated the effect of external divalent cations on pacemaking activity in cultured ICC from murine small intestine by using whole-cell patch clamp techniques. ICC generated pacemaker currents under a voltage clamp or electrical pacemaker potentials under a current clamp, and showed a mean amplitude of $-500{\pm}50$ pA or $30{\pm}1$ mV and the frequency of $18{\pm}2$ cycles/min. Treatments of the cells with external 0 mM $Ca^{2+}$ stopped pacemaking activity of ICC. In the presence of 2 mM $Ca^{2+}$, 0 mM external $Mg^{2+}$ depolarized the resting membrane potential, and there was no change in the frequency of pacemaking activity. However, 10 mM external $Mg^{2+}$ decreased the frequency of pacemaking activity ($6.75{\pm}1$ cycles/min, n=5). We replaced external 2 mM $Ca^{2+}$ with equimolar $Ba^{2+}$, $Mn^{2+}$ and $Sr^{2+}$, and they all developed inward current in the sequence of $Ba^{2+}$>$Mn^{2+}$>$Sr^{2+}$. Also the frequency of the pacemaking activity was stopped or irregulated. We investigated the effect of 10 mM $Ba^{2+}$, $Mn^{2+}$ and $Sr^{2+}$ on pacemaking activity of ICC in the presence of external 0 mM $Mg^{2+}$, and found that 10 mM $Ba^{2+}$ and $Mn^{2+}$ induced large inward current and stopped the pacemaking activity of ICC (n=5). Interestingly, 10 mM $Sr^{2+}$ induced small inward current and potentiated the amplitude of pacemaking activity of ICC (n=5). These results indicate that extracellular $Ca^{2+}$ and $Mg^{2+}$ are requisite for the pacemaking activity of ICC.

• 한국응용약물학회:학술대회논문집
• /
• 한국응용약물학회 1994년도 춘계학술대회 and 제3회 신약개발 연구발표회
• /
• pp.217-217
• /
• 1994

Calcium entry blockers, capable of inhibiting transmembrane influx of extracellular calcium through specific calcium channels, are useful drugs in the treatment of angina pectoris, hypertension, cardiac arrythmia, and various cardiovascular disorders. Compounds having isoquinoline structures have recently been reported to possess calcium antagonistic action. Therefore, in the present study, we have attempted to synthesize some isoquinoline and related compound.; in order to search for potentially effective chemicals acting on cardiovascular system, and evaluated their pharmacological properties focusing on calcium antagonistic actions. Almost all of the compounds so far synthesized, had inhibitory action against phenylephrine or high potassium-induced contraction in vascular smooth muscle with different degrees of potencies depending on their structures, However, some of tetrahydroisoquinoline analogs showed directly inhibit calcium current in isolated rabbit cardiac myocytes examined by patch clamp techniques. The pharmacological properties of these compounds need more intensive investigation as to whether these chemicals may have developed as a new cardiovascular active drugs. Therefore, we are now under investigation of the mechanism of action of these compounds.

• The Korean Journal of Physiology and Pharmacology
• /
• 제4권2호
• /
• pp.81-90
• /
• 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.