Vasorelaxing Activity of R-(−)-3′-Hydroxy-2,4,5-trimethoxydalbergiquinol from Dalbergia tonkinensis: Involvement of Smooth Muscle CaV1.2 Channels

Planta Medica ◽  
2020 ◽  
Vol 86 (04) ◽  
pp. 284-293 ◽  
Author(s):  
Nguyen Manh Cuong ◽  
Ninh The Son ◽  
Ngu Truong Nhan ◽  
Pham Ngoc Khanh ◽  
Tran Thu Huong ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Antihypertensive Drugs ◽  
Docking Simulation ◽  
Mechanical Activity ◽  
Dependent Manner ◽  
Vascular Effects ◽  
Patch Clamp Technique ◽  
Spasmolytic Activity ◽  
Cav1.2 Channels ◽  
Dalbergia Tonkinensis

Abstract Dalbergia species heartwood, widely used in traditional medicine to treat various cardiovascular diseases, might represent a rich source of vasoactive agents. In Vietnam, Dalbergia tonkinensis is an endemic tree. Therefore, the aim of the present work was to investigate the vascular activity of R-(−)-3′-hydroxy-2,4,5-trimethoxydalbergiquinol isolated from the heartwood of D. tonkinensis and to provide circular dichroism features of its R absolute configuration. The vascular effects of R-(−)-3′-hydroxy-2,4,5-trimethoxydalbergiquinol were assessed on the in vitro mechanical activity of rat aorta rings, under isometric conditions, and on whole-cell Ba2+ currents through CaV1.2 channels (IBa1.2) recorded in single, rat tail main artery myocytes by means of the patch-clamp technique. R-(−)-3′-Hydroxy-2,4,5-trimethoxydalbergiquinol showed concentration-dependent, vasorelaxant activity on both endothelium-deprived and endothelium intact rings precontracted with the α 1 receptor agonist phenylephrine. Neither the NO (nitric oxide) synthase inhibitor Nω-nitro-L-arginine methyl ester nor the cyclooxygenase inhibitor indomethacin affected its spasmolytic activity. R-(−)-3′-Hydroxy-2,4,5-trimethoxydalbergiquinol-induced vasorelaxation was antagonized by (S)-(−)-Bay K 8644 and unaffected by tetraethylammonium plus glibenclamide. In patch-clamp experiments, R-(−)-3′-hydroxy-2,4,5-trimethoxydalbergiquinol inhibited IBa1.2 in a concentration-dependent manner and significantly decreased the time constant of current inactivation. R-(−)-3′-Hydroxy-2,4,5-trimethoxydalbergiquinol likely stabilized the channel in its closed state, as suggested by molecular modelling and docking simulation to the CaV1.2 channel α 1c subunit. In conclusion, D. tonkinensis species may represent a source of agents potentially useful for the development of novel antihypertensive drugs.

Vasorelaxing Activity of Stilbenoid and Phenanthrene Derivatives from Brasiliorchis porphyrostele: Involvement of Smooth Muscle CaV1.2 Channels

Planta Medica ◽  
2020 ◽  
Vol 86 (09) ◽  
pp. 631-642
Author(s):  
Watcharee Waratchareeyakul ◽  
Fabio Fusi ◽  
Miriam Durante ◽  
Amer Ahmed ◽  
Walter Knirsch ◽  
...  
Keyword(s):  
Channel Blocker ◽  
Antihypertensive Agents ◽  
Dependent Manner ◽  
Vascular Effects ◽  
Concentration Dependent Manner ◽  
Spasmolytic Activity ◽  
Depth Analysis ◽  
Cav1.2 Channels ◽  
Vasorelaxant Activity

AbstractFive compounds, 3,4′-dihydroxy-3′,5,5′-trimethoxydihydrostilbene, 1; 3,4′-ihydroxy-3′,5′-dimethoxydihydrostilbene, 2; 3,4′-dihydroxy-5,5′-dimethoxydihydrostilbene, 3; 9,10-dihydro-2,7-dihydroxy-4,6-dimethoxyphenanthrene, 4; and the previously unreported 1,2,6,7-tetrahydroxy-4-methoxyphenanthrene, 5 were isolated from the South American orchid, Brasiliorchis porphyrostele. An in-depth analysis of their vascular effects was performed on in vitro rat aorta rings and tail main artery myocytes. Compounds 1 – 4 were shown to possess vasorelaxant activity on rings pre-contracted by the α 1 receptor agonist phenylephrine, the CaV1.2 stimulator (S)-(−)-Bay K 8644, or depolarized with high K+ concentrations. However, compound 5 was active solely on rings stimulated by 25 mM but not 60 mM K+. The spasmolytic activity of compounds 1 and 4 was significantly affected by the presence of an intact endothelium. The KATP channel blocker glibenclamide and the KV channel blocker 4-aminopyridine significantly antagonized the vasorelaxant activity of compounds 4 and 1, respectively. In patch-clamp experiments, compounds 1 – 4 inhibited Ba2+ current through CaV1.2 channels in a concentration-dependent manner, whereas neither compound 4 nor compound 1 affected K+ currents through KATP and KV channels, respectively. The present in vitro, comprehensive study demonstrates that Brasiliorchis porphyrostele may represent a source of vasoactive agents potentially useful for the development of novel antihypertensive agents that has now to be validated in vivo in animal models of hypertension.

Gramicidin perforated patch-clamp technique reveals glycine-gated outward chloride current in dissociated nucleus solitarii neurons of the rat

1994 ◽  
Vol 72 (3) ◽  
pp. 1103-1108 ◽  
Author(s):  
J. S. Rhee ◽  
S. Ebihara ◽  
N. Akaike
Keyword(s):  
Patch Clamp ◽  
Hill Coefficient ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Perforated Patch ◽  
Clamp Technique ◽  
Outward Currents ◽  
Patch Technique ◽  
Perforated Patch Clamp

1. The inhibitory response of exogenously applied glycine was investigated in freshly dissociated rat nucleus tractus solitarii neurons under whole cell configuration using new perforated patch-clamp technique termed "gramicidin perforated patch technique," which maintains intact intracellular Cl- concentrations. 2. Using the gramicidin perforated patch technique, at a holding potential (VH) of -45 mV, glycine induced outward currents in a concentration-dependent manner with a EC50 of 4.0 x 10(-5) M and at a Hill coefficient of 1.5. In contrast, using the nystatin perforated patch technique, glycine induced inward currents at the same VH in a concentration-dependent manner with an EC50 of 4.9 x 10(-5) M and at a Hill coefficient of 1.2. 3. The glycine-induced outward currents were blocked by strychnine in a concentration dependent manner with an IC50 of 2.2 x 10(-8) M. The blockade was competitive. 4. The current-voltage relationship for the 10(-5) M glycine response showed a clear outward rectification. 5. Ten-fold change of extracellular Cl- with a large impermeable anion resulted in a 65 mV shift of the reversal potential of glycine-induced currents (EGly), indicating that the membrane behaves like a Cl- electrode in the presence of glycine. 6. The intracellular Cl- activity calculated from the EGly ranged from 7.3 to 18.2 mM, with a mean value of 13.3 mM. 7. The values of EGly in the individual neurons were significantly negative to the resting membrane potentials, suggesting the existence of active transport of Cl-.

Effects of the gap junction blocker glycyrrhetinic acid on gastrointestinal smooth muscle cells

2005 ◽  
Vol 288 (4) ◽  
pp. G832-G841 ◽  
Author(s):  
Yukari Takeda ◽  
Sean M. Ward ◽  
Kenton M. Sanders ◽  
Sang Don Koh
Keyword(s):  
Smooth Muscle ◽  
Patch Clamp ◽  
Smooth Muscle Cells ◽  
Gap Junctions ◽  
Lucifer Yellow ◽  
Muscle Cells ◽  
Glycyrrhetinic Acid ◽  
Delayed Rectifier ◽  
Dependent Manner ◽  
Patch Clamp Technique

In the tunica muscularis of the gastrointestinal (GI) tract, gap junctions form low-resistance pathways between pacemaker cells known as interstitial cells of Cajal (ICCs) and between ICC and smooth muscle cells. Coupling via these junctions facilitates electrical slow-wave propagation and responses of smooth muscle to enteric motor nerves. Glycyrrhetinic acid (GA) has been shown to uncouple gap junctions, but previous studies have shown apparent nonspecific effects of GA in a variety of tissues. We tested the effects of GA using isometric force measurements, intracellular microelectrode recordings, the patch-clamp technique, and the spread of Lucifer yellow within cultured ICC networks. In murine small intestinal muscles, β-GA (10 μM) decreased phasic contractions and depolarized resting membrane potential. Preincubation of GA inhibited the spread of Lucifer yellow, increased input resistance, and decreased cell capacitance in ICC networks, suggesting that GA uncoupled ICCs. In patch-clamp experiments of isolated jejunal myocytes, GA significantly decreased L-type Ca2+ current in a dose-dependent manner without affecting the voltage dependence of this current. The IC50 for Ca2+ currents was 1.9 μM, which is lower than the concentrations used to block gap junctions. GA also significantly increased large-conductance Ca2+-activated K+ currents but decreased net delayed rectifier K+ currents, including 4-aminopyridine and tetraethylammonium-resistant currents. In conclusion, the reduction of phasic contractile activity of GI muscles by GA is likely a consequence of its inhibitory effects on gap junctions and voltage-dependent Ca2+ currents. Membrane depolarization may be a consequence of uncoupling effects of GA on gap junctions between ICCs and smooth muscles and inhibition of K+ conductances in smooth muscle cells.

Characterizations of the Urate Transporter, GLUT9, and Its Potent Inhibitors by Patch-Clamp Technique

2020 ◽  
pp. 247255522094950
Author(s):  
Yanyu Chen ◽  
Zean Zhao ◽  
Yongmei Li ◽  
Lu Li ◽  
Yu Jiang ◽  
...  
Keyword(s):  
Uric Acid ◽  
Patch Clamp ◽  
Glucose Transporter ◽  
Cell Model ◽  
Outward Current ◽  
External Solution ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Clamp Technique ◽  
Induced Currents

Glucose transporter 9 (GLUT9), which transports urate in an electrogenic and voltage-dependent manner, plays an important role in the maintenance of normal blood uric acid/urate levels. In the present study, we established a cell model based on the single-electrode patch-clamp technique for characterization of GLUT9 and explored the inhibitory effects of benzobromarone (BM) and probenecid (PB) on urate-induced currents in mouse GLUT9a (mGLUT9a)–expressing HEK-293T cells. The results showed that uric acid, rather than glucose perfusion, led to a rapid and large outward current by mGLUT9a in dose-, voltage-, and pH-dependent manners. BM prominently and irreversibly inhibited the uric acid–induced currents through mGLUT9a, and PB weakly and reversibly inhibited mGLUT9a. We found that depletion of K+ in the external solution significantly strengthened the blockade of BM on mGLUT9a. In addition, an enhanced inhibitory rate of BM was detected when the pH of the external solution was changed from 7.4 to 5.5, indicating that BM functions optimally in an acidic environment. In conclusion, the combination of the established cell model with patch-clamp techniques first revealed the function properties of GLUT9 inhibitors and may provide potential benefits to the study of GLUT9 inhibitors as antihyperuricemic or antigout agents.

Dendritic excitability and a voltage-gated calcium current in locust nonspiking local interneurons

1993 ◽  
Vol 69 (5) ◽  
pp. 1484-1498 ◽  
Author(s):  
G. Laurent ◽  
K. J. Seymour-Laurent ◽  
K. Johnson
Keyword(s):  
Patch Clamp ◽  
Resting Potential ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Inward Current ◽  
Time To Peak ◽  
Clamp Technique ◽  
Voltage Dependent

1. The active properties of axonless nonspiking interneurons in the thoracic nervous system of the locust Schistocerca americana were studied in vivo with the switched current-clamp technique from dendritic impalements, and in vitro with the whole-cell variation of the patch-clamp technique. 2. In 20% of in vivo recordings, depolarization of a dendrite to potentials more positive than about -40 mV evoked resonant behaviour and/or regenerative potentials. The latter were slow (half width: 20-30 ms), small (base-to-peak amplitude: 25-35 mV), and were often followed by a pronounced after hyperpolarization (AHP). 3. The slow regenerative potentials sometimes had multiple peaks separated by incomplete repolarizations. The voltage envelope of such potentials was always broader than that of spikes with single peaks. In other recordings, a same depolarizing pulse could evoke several regenerative potentials with different waveforms. These results suggested the presence of multiple dendritic initiation sites separated by regions of inexcitable membrane, allowing decremental conduction and the passive fusion of spike envelopes. 4. Graded active responses could also be evoked on rebound from short hyperpolarizations such as inhibitory postsynaptic potentials (IPSPs) provided that the membrane was already depolarized to about -40 mV. IPSPs evoked by several presynaptic interneurons differed in their ability to evoke rebound potentials suggesting that some synaptic sites were electrically closer than others to regions of active membrane. 5. Patch-clamp recordings from somata of nonspiking neurons isolated from 75% embryos and grown in culture medium for 1-2 days revealed the presence of an inactivating inward current resistant to 0.5-1 microM tetrodotoxin (TTX). The inward current was carried equally well by Ba2+, and sensitive to blockade by Cd2+ (0.5 mM), Ni2+ (0.75 mM), or Co2+ (2.5 mM). 6. The current activated around -40 mV, with voltage-dependent activation (time-to-peak approximately 20 ms at -35 mV and 1-2 ms at 0 mV). Tail currents evoked upon repolarization were well fitted by a single exponential (tau = 1-2 ms). Deactivation time constants shorter than 300 microseconds, however, could not be measured. 7. The current inactivated rapidly in a voltage-dependent manner, following two-exponential kinetics. A very small persistent component could be explained by the overlap between activation and inactivation curves, greatest at approximately -20 mV. The voltage of half-inactivation was about -25 mV. At a resting potential of -58 mV, 90% of the current was available for activation. Recovery from steady-state inactivation followed the sum of two or more exponential processes.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals A patch-clamp study of histamine-secreting cells.

1986 ◽  
Vol 88 (3) ◽  
pp. 349-368 ◽  
Author(s):  
M Lindau ◽  
J M Fernandez
Keyword(s):  
Mast Cells ◽  
Patch Clamp ◽  
Single Channel ◽  
Resting Potential ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Peritoneal Mast Cells ◽  
Ionic Conductances ◽  
Rat Peritoneal Mast Cells

The ionic conductances in rat basophilic leukemia cells (RBL-2H3) and rat peritoneal mast cells were investigated using the patch-clamp technique. These two cell types were found to have different electrophysiological properties in the resting state. The only significant conductance of RBL-2H3 cells was a K+-selective inward rectifier. The single channel conductance at room temperature increased from 2-3 pS at 2.8 mM external K+ to 26 pS at 130 mM K+. This conductance, which appeared to determine the resting potential, could be blocked by Na+ and Ba2+ in a voltage-dependent manner. Rat peritoneal mast cells had a whole-cell conductance of only 10-30 pS, and the resting potential was close to zero. Sometimes discrete openings of channels were observed in the whole-cell configuration. When the Ca2+ concentration on the cytoplasmic side of the membrane was elevated, two types of channels with poor ion specificity appeared. A cation channel, observed at a Ca2+ concentration of approximately 1 microM, had a unit conductance of 30 pS. The other channel, activated at several hundred micromolar Ca2+, was anion selective and had a unit conductance of approximately 380 pS in normal Ringer solution and a bell-shaped voltage dependence. Antigenic stimulation did not cause significant changes in the ionic conductances in either cell type, which suggests that these cells use a mechanism different from ionic currents in stimulus-secretion coupling.

scholarly journals Inhibition of cardiac Na+ currents by isoproterenol

1990 ◽  
Vol 258 (4) ◽  
pp. H977-H982 ◽  
Author(s):  
B. Schubert ◽  
A. M. Vandongen ◽  
G. E. Kirsch ◽  
A. M. Brown
Keyword(s):  
Patch Clamp ◽  
Ventricular Myocytes ◽  
Neonatal Rat ◽  
Na Channel ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Whole Cell Patch Clamp ◽  
Na Currents ◽  
Voltage Dependent

The mechanism by which the beta-adrenergic agonist isoproterenol (ISO) modulates voltage-dependent cardiac Na+ currents (INa) was studied in single ventricular myocytes of neonatal rat using the gigaseal patch-clamp technique. ISO inhibited INa reversibly, making the effect readily distinguishable from the monotonic decrease of INa caused by the shift in gating that customarily occurs during whole cell patch-clamp experiments (E. Fenwick, A. Marty, and E. Neher, J. Physiol. Lond. 331: 599-635, 1982; and J. M. Fernandez, A. P. Fox, and S. Krasne, J. Physiol. Lond. 356: 565-585, 1984). The inhibition was biphasic, having fast and slow components, and was voltage-dependent, being more pronounced at depolarized potentials. In whole cell experiments the membrane-permeable adenosine 3',5'-cyclic monophosphate (cAMP) congener 8-bromo-cAMP reduced INa. In cell-free inside-out patches with ISO present in the pipette, guanosine 5'-triphosphate (GTP) applied to the inner side of the membrane patch inhibited single Na+ channel activity. This inhibition could be partly reversed by hyperpolarizing prepulses. The nonhydrolyzable GTP analogue guanosine-5'-O-(3-thiotriphosphate) greatly reduced the probability of single Na+ channel currents in a Mg2(+)-dependent manner. We propose that ISO inhibits cardiac Na+ channels via the guanine nucleotide binding, signal-transducing G protein that acts through both direct (membrane delimited) and indirect (cytoplasmic) pathways.

Neural effects of parathyroid hormone: modulation of the calcium channel current and metabolism of monoamines in identified Helisoma snail neurons

1993 ◽  
Vol 71 (8) ◽  
pp. 582-591 ◽  
Author(s):  
R. Wang ◽  
P. K. T. Pang ◽  
L. Wu ◽  
A. Shipley ◽  
E. Karpinski ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Patch Clamp ◽  
Calcium Channel ◽  
Dependent Manner ◽  
Vibrating Probe ◽  
Buccal Ganglion ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Buccal Ganglia ◽  
Channel Currents

Neuronal effects of parathyroid hormone (PTH) have been reported in vertebrates. The effect of PTH on invertebrate central neurons within the buccal ganglion of Helisoma trivolvis snails was examined in the present study. By using a vibrating probe, PTH was found to induce a transient calcium-dependent inward current in intact buccal ganglia. Intracellular micro-electrode recording revealed that PTH broadened the spontaneous action potential in buccal B5 neurons in situ. By using the whole-cell configuration of the patch-clamp technique, PTH was demonstrated to increase the N-like calcium channel currents in isolated B5 neurons in a concentration-dependent manner. This effect of PTH on the N-like calcium channel currents depended on the activation of a G protein insensitive to pertussis toxin, but was unlikely to be mediated by the cyclic AMP dependent protein kinase. Furthermore, the release of γ-glutamyl conjugate of dopamine from buccal ganglia was selectively increased in the presence of PTH. These results represent the first demonstration that a vertebrate peptide hormone, PTH, selectively modulates the N-like voltage-dependent calcium channel currents in identified invertebrate neurons. Therefore, a novel role of PTH in the regulation of invertebrate central neural functions is indicated.Key words: parathyroid hormone, patch clamp, vibrating probe, dopamine, invertebrate neuron.

scholarly journals Oxytocin Inhibits T-Type Calcium Current of Human Decidual Stromal Cells

2005 ◽  
Vol 90 (7) ◽  
pp. 4191-4197 ◽  
Author(s):  
Bo Liu ◽  
Stephen J. Hill ◽  
Raheela N. Khan
Keyword(s):  
Stromal Cells ◽  
Calcium Current ◽  
Elective Cesarean Section ◽  
Dependent Manner ◽  
Uterine Contractility ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Inactivation Curve ◽  
Decidual Cells ◽  
Elective Cesarean

Abstract Context: Little is known about the crosstalk between the decidua and myometrium in relation to human labor. The hormone oxytocin (OT) is considered to be a key mediator of uterine contractility during parturition, exerting some of its effects through calcium channels. Objective: The objective was to characterize the effect of OT on the T-type calcium channel in human decidual stromal cells before and after the onset of labor. Design: The nystatin-perforated patch-clamp technique was used to record inward T-type calcium current (ICa(T)) from acutely dispersed decidual stromal cells obtained from women at either elective cesarean section [CS (nonlabor)] or after normal spontaneous vaginal delivery [SVD (labor)]. Setting: These studies took place at the University of Nottingham Medical School. Results: I Ca(T) of both SVD and CS cells were blocked by nickel (IC50 of 5.6 μm) and cobalt chloride (1 mm) but unaffected by nifedipine (10 μm). OT (1 nm to 3.5 μm) inhibited ICa(T) of SVD cells in a concentration-dependent manner, with a maximal inhibition of 79.0% compared with 26.2% in decidual cells of the CS group. OT-evoked reduction of ICa(T) was prevented by preincubation with the OT antagonist L371,257 in the SVD but not CS group. OT, in a concentration-dependent manner, displaced the steady-state inactivation curve for ICa(T) to the left in the SVD group with no significant effect on curves of the CS group. Conclusion: Inhibition of ICa(T) by OT in decidual cells obtained during labor may signify important functional remodeling of uterine signaling during this period.

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