scholarly journals The Effectiveness in Activating M-Type K+ Current Produced by Solifenacin ([(3R)-1-azabicyclo[2.2.2]octan-3-yl] (1S)-1-phenyl-3,4-dihydro-1H-isoquinoline-2-carboxylate): Independent of Its Antimuscarinic Action

2021 ◽  
Vol 22 (22) ◽  
pp. 12399
Author(s):  
Hsin-Yen Cho ◽  
Tzu-Hsien Chuang ◽  
Sheng-Nan Wu
Keyword(s):  
Single Channel ◽  
Ionic Currents ◽  
Reaction Scheme ◽  
Gh3 Cells ◽  
Whole Cell ◽  
Type K ◽  
Cell Current ◽  
K Current ◽  
Subsequent Addition ◽  
Whole Cell Current

Solifenacin (Vesicare®, SOL), known to be a member of isoquinolines, is a muscarinic antagonist that has anticholinergic effect, and it has been beneficial in treating urinary incontinence and neurogenic detrusor overactivity. However, the information regarding the effects of SOL on membrane ionic currents is largely uncertain, despite its clinically wide use in patients with those disorders. In this study, the whole-cell current recordings revealed that upon membrane depolarization in pituitary GH3 cells, the exposure to SOL concentration-dependently increased the amplitude of M-type K+ current (IK(M)) with effective EC50 value of 0.34 μM. The activation time constant of IK(M) was concurrently shortened in the SOL presence, hence yielding the KD value of 0.55 μM based on minimal reaction scheme. As cells were exposed to SOL, the steady-state activation curve of IK(M) was shifted along the voltage axis to the left with no change in the gating charge of the current. Upon an isosceles-triangular ramp pulse, the hysteretic area of IK(M) was increased by adding SOL. As cells were continually exposed to SOL, further application of acetylcholine (1 μM) failed to modify SOL-stimulated IK(M); however, subsequent addition of thyrotropin releasing hormone (TRH, 1 μM) was able to counteract SOL-induced increase in IK(M) amplitude. In cell-attached single-channel current recordings, bath addition of SOL led to an increase in the activity of M-type K+ (KM) channels with no change in the single channel conductance; the mean open time of the channel became lengthened. In whole-cell current-clamp recordings, the SOL application reduced the firing of action potentials (APs) in GH3 cells; however, either subsequent addition of TRH or linopirdine was able to reverse SOL-mediated decrease in AP firing. In hippocampal mHippoE-14 neurons, the IK(M) was also stimulated by adding SOL. Altogether, findings from this study disclosed for the first time the effectiveness of SOL in interacting with KM channels and hence in stimulating IK(M) in electrically excitable cells, and this noticeable action appears to be independent of its antagonistic activity on the canonical binding to muscarinic receptors expressed in GH3 or mHippoE-14 cells.

scholarly journals Characterization of Effectiveness in Concerted Ih Inhibition and IK(Ca) Stimulation by Pterostilbene (Trans-3,5-dimethoxy-4′-hydroxystilbene), a Stilbenoid

2020 ◽  
Vol 21 (1) ◽  
pp. 357 ◽  
Author(s):  
Edmund Cheung So ◽  
Zi-Han Gao ◽  
Shun Yao Ko ◽  
Sheng-Nan Wu
Keyword(s):  
Single Channel ◽  
Ionic Currents ◽  
Gh3 Cells ◽  
Dependent Manner ◽  
Excitable Cells ◽  
Bkca Channels ◽  
Concentration Dependent Manner ◽  
Cell Current ◽  
K Current ◽  
Neuroprotective Actions

Pterostilbene (PTER), a natural dimethylated analog of resveratrol, has been demonstrated to produce anti-neoplastic or neuroprotective actions. However, how and whether this compound can entail any perturbations on ionic currents in electrically excitable cells remains unknown. In whole-cell current recordings, addition of PTER decreased the amplitude of macroscopic Ih during long-lasting hyperpolarization in GH3 cells in a concentration-dependent manner, with an effective IC50 value of 0.84 μM. Its presence also shifted the activation curve of Ih along the voltage axis to a more hyperpolarized potential, by 11 mV. PTER at a concentration greater than 10 μM could also suppress l-type Ca2+ and transient outward K+ currents in GH3 cells. With the addition of PTER, IK(Ca) amplitude was increased, with an EC50 value of 2.23 μM. This increase in IK(Ca) amplitude was attenuated by further addition of verruculogen, but not by tolbutamide or TRAM-39. Neither atropine nor nicotine, in the continued presence of PTER, modified the PTER-stimulated IK(Ca). PTER (10 μM) slightly suppressed the amplitude of l-type Ca2+ current and transient outward K+ current. The presence of PTER (3 μM) was also effective at increasing the open-state probability of large-conductance Ca2+-activated K+ (BKCa) channels identified in hippocampal mHippoE-14 neurons; however, its inability to alter single-channel conductance was detected. Our study highlights evidence to show that PTER has the propensity to perturb ionic currents (e.g., Ih and IK(Ca)), thereby influencing the functional activities of neurons, and neuroendocrine or endocrine cells.

scholarly journals Volume-Dependent Atp-Conductive Large-Conductance Anion Channel as a Pathway for Swelling-Induced Atp Release

2001 ◽  
Vol 118 (3) ◽  
pp. 251-266 ◽  
Author(s):  
Ravshan Z. Sabirov ◽  
Amal K. Dutta ◽  
Yasunobu Okada
Keyword(s):  
Single Channel ◽  
Atp Release ◽  
Anion Channel ◽  
Time Dependent ◽  
Whole Cell ◽  
Dependent Inactivation ◽  
Current Voltage ◽  
Voltage Dependent ◽  
Cell Current ◽  
Whole Cell Current

In mouse mammary C127i cells, during whole-cell clamp, osmotic cell swelling activated an anion channel current, when the phloretin-sensitive, volume-activated outwardly rectifying Cl− channel was eliminated. This current exhibited time-dependent inactivation at positive and negative voltages greater than around ±25 mV. The whole-cell current was selective for anions and sensitive to Gd3+. In on-cell patches, single-channel events appeared with a lag period of ∼15 min after a hypotonic challenge. Under isotonic conditions, cell-attached patches were silent, but patch excision led to activation of currents that consisted of multiple large-conductance unitary steps. The current displayed voltage- and time-dependent inactivation similar to that of whole-cell current. Voltage-dependent activation profile was bell-shaped with the maximum open probability at −20 to 0 mV. The channel in inside-out patches had the unitary conductance of ∼400 pS, a linear current-voltage relationship, and anion selectivity. The outward (but not inward) single-channel conductance was suppressed by extracellular ATP with an IC50 of 12.3 mM and an electric distance (δ) of 0.47, whereas the inward (but not outward) conductance was inhibited by intracellular ATP with an IC50 of 12.9 mM and δ of 0.40. Despite the open channel block by ATP, the channel was ATP-conductive with PATP/PCl of 0.09. The single-channel activity was sensitive to Gd3+, SITS, and NPPB, but insensitive to phloretin, niflumic acid, and glibenclamide. The same pharmacological pattern was found in swelling-induced ATP release. Thus, it is concluded that the volume- and voltage-dependent ATP-conductive large-conductance anion channel serves as a conductive pathway for the swelling-induced ATP release in C127i cells.

scholarly journals Endogenous chloride channels of insect sf9 cells. Evidence for coordinated activity of small elementary channel units.

1996 ◽  
Vol 107 (6) ◽  
pp. 695-714 ◽  
Author(s):  
E H Larsen ◽  
S E Gabriei ◽  
M J Stutts ◽  
J Fullton ◽  
E M Price ◽  
...  
Keyword(s):  
Chloride Channels ◽  
Single Channel ◽  
Low Frequency ◽  
Power Density Spectrum ◽  
Current Fluctuations ◽  
Sf9 Cells ◽  
Density Spectrum ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

The endogenous Cl- conductance of Spodoptera frugiperda (Sf9) cells was studied 20-35 h after plating out of either uninfected cells or cells infected by a baculovirus vector carrying the cloned beta-galactosidase gene (beta-Gal cells). With the cation Tris+ in the pipette and Na+ in the bath, the reversal potential of whole-cell currents was governed by the prevailing Cl- equilibrium potential and could be fitted by the Goldman-Hodgkin-Katz equation with similar permeabilities for uninfected and beta-Gal cells. In the frequency range 0.12 < f < 300 Hz, the power density spectrum of whole-cell Cl- currents could be fitted by three Lorentzians. Independent of membrane potential, >50% of the total variance of whole-cell current fluctuations was accounted for by the low frequency Lorentzian (fc = 0.40 +/- 0.03 Hz, n = 6). Single-Cl- channels showed complex gating kinetics with long lasting (seconds) openings interrupted by similar long closures. In the open state, channels exhibited fast burst-like closures. Since the patches normally contained more than a single channel, it was not possible to measure open and closed dwell-time distributions for comparing single-Cl- channel activity with the kinetic features of whole-cell currents. However, the power density spectrum of Cl- currents of cell-attached and excised outside-out patches contained both high and low frequency Lorentzian components, with the corner frequency of the slow component (fc = 0.40 +/- 0.02 Hz, n = 4) similar to that of whole-cell current fluctuations. Chloride channels exhibited multiple conductance states with similar Goldman-Hodgkin-Katz-type rectification. Single-channel permeabilities covered the range from approximately 0.6.10(-14) cm5/s to approximately 6.10(-14) cm3/s, corresponding to a limiting conductance (gamma 150/150) of approximately 3.5 pS and approximately 35 pS, respectively. All states reversed near the same membrane potential, and they exhibited similar halide ion selectivity, P1 > PCl approximately PBr. Accordingly, Cl- current amplitudes larger than current flow through the smallest channel unit resolved seem to result from simultaneous open/shut events of two or more channel units.

scholarly journals Cell swelling activates ATP-dependent voltage-gated chloride channels in M-1 mouse cortical collecting duct cells.

1996 ◽  
Vol 108 (3) ◽  
pp. 177-193 ◽  
Author(s):  
K Meyer ◽  
C Korbmacher
Keyword(s):  
Voltage Dependence ◽  
Single Channel ◽  
Collecting Duct ◽  
High Energy ◽  
Cell Swelling ◽  
Single Channels ◽  
Cortical Collecting Duct ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

In the present study we used whole-cell patch clamp recordings to investigate swelling-activated Cl-currents (ICl-swell) in M-1 mouse cortical collecting duct (CCD) cells. Hypotonic cell swelling reversibly increased the whole-cell Cl- conductance by about 30-fold. The I-V relationship was outwardly-rectifying and ICl-swell displayed a characteristic voltage-dependence with relatively fast inactivation upon large depolarizing and slow activation upon hyperpolarizing voltage steps. Reversal potential measurements revealed a selectivity sequence SCN- > I- > Br- > Cl- > > gluconate. ICl-swell was inhibited by tamoxifen, NPPB (5-nitro-2(3-phenylpropylamino)-benzoate), DIDS (4,4'-diisothiocyanostilbene-2,2'-disulphonic acid), flufenamic acid, niflumic acid, and glibenclamide, in descending order of potency. Extracellular cAMP had no significant effect. ICl-swell was Ca2+ independent, but current activation depended on the presence of a high-energy gamma-phosphate group from intracellular ATP or ATP gamma S. Moreover, it depended on the presence of intracellular Mg2+ and was inhibited by staurosporine, which indicates that a phosphorylation step is involved in channel activation. Increasing the cytosolic Ca2+ concentration by using ionomycin stimulated Cl- currents with a voltage dependence different from that of ICl-swell. Analysis of whole-cell current records during early onset of ICl-swell and during final recovery revealed discontinuous step-like changes of the whole-cell current level which were not observed under nonswelling conditions. A single-channel I-V curve constructed using the smallest resolvable current transitions detected at various holding potentials and revealed a slope conductance of 55, 15, and 8 pS at +120, 0, and -120 mV, respectively. The larger current steps observed in these recordings had about 2, 3, or 4 times the size of the putative single-channel current amplitude, suggesting a coordinated gating of several individual channels or channel subunits. In conclusion we have functionally characterized ICl-swell in M-1 CCD cells and have identified the underlying single channels in whole-cell current recordings.

scholarly journals Effective Activation by Kynurenic Acid and Its Aminoalkylated Derivatives on M-Type K+ Current

2021 ◽  
Vol 22 (3) ◽  
pp. 1300
Author(s):  
Yi-Ching Lo ◽  
Chih-Lung Lin ◽  
Wei-Yu Fang ◽  
Bálint Lőrinczi ◽  
István Szatmári ◽  
...  
Keyword(s):  
Kynurenic Acid ◽  
Time Course ◽  
Ionic Currents ◽  
Gh3 Cells ◽  
Excitable Cells ◽  
Patch Clamp Technique ◽  
Open Probability ◽  
Type K ◽  
K Current

Kynurenic acid (KYNA, 4-oxoquinoline-2-carboxylic acid), an intermediate of the tryptophan metabolism, has been recognized to exert different neuroactive actions; however, the need of how it or its aminoalkylated amide derivative N-(2-(dimethylamino)ethyl)-3-(morpholinomethyl)-4-oxo-1,4-dihydroquinoline-2-carboxamide (KYNA-A4) exerts any effects on ion currents in excitable cells remains largely unmet. In this study, the investigations of how KYNA and other structurally similar KYNA derivatives have any adjustments on different ionic currents in pituitary GH3 cells and hippocampal mHippoE-14 neurons were performed by patch-clamp technique. KYNA or KYNA-A4 increased the amplitude of M-type K+ current (IK(M)) and concomitantly enhanced the activation time course of the current. The EC50 value required for KYNA- or KYNA-A4 -stimulated IK(M) was yielded to be 18.1 or 6.4 μM, respectively. The presence of KYNA or KYNA-A4 shifted the relationship of normalized IK(M)-conductance versus membrane potential to more depolarized potential with no change in the gating charge of the current. The voltage-dependent hysteretic area of IK(M) elicited by long-lasting triangular ramp pulse was observed in GH3 cells and that was increased during exposure to KYNA or KYNA-A4. In cell-attached current recordings, addition of KYNA raised the open probability of M-type K+ channels, along with increased mean open time of the channel. Cell exposure to KYNA or KYNA-A4 mildly inhibited delayed-rectifying K+ current; however, neither erg-mediated K+ current, hyperpolarization-activated cation current, nor voltage-gated Na+ current in GH3 cells was changed by KYNA or KYNA-A4. Under whole-cell, current-clamp recordings, exposure to KYNA or KYNA-A4 diminished the frequency of spontaneous action potentials; moreover, their reduction in firing frequency was attenuated by linopirdine, yet not by iberiotoxin or apamin. In hippocampal mHippoE-14 neurons, the addition of KYNA also increased the IK(M) amplitude effectively. Taken together, the actions presented herein would be one of the noticeable mechanisms through which they modulate functional activities of excitable cells occurring in vivo.

Effect of volume changes on a potassium current in rabbit corneal epithelial cells

1993 ◽  
Vol 264 (5) ◽  
pp. C1238-C1245 ◽  
Author(s):  
G. Farrugia ◽  
J. Rae
Keyword(s):  
Epithelial Cells ◽  
Chloride Ion ◽  
Ion Concentration ◽  
K Channel ◽  
Corneal Epithelial Cells ◽  
Whole Cell ◽  
Corneal Epithelial ◽  
Cell Current ◽  
K Current ◽  
Whole Cell Current

Patch-clamp recordings from rabbit corneal epithelial cells have identified a large-conductance (167 pS in symmetrical 150 mM KCl) K channel that is the major contributor to the whole cell current (J. L. Rae and G. Farrugia. J. Membr. Biol. 129: 81-87, 1992). We report here on the regulation of this channel by changes in cellular osmolality and/or volume. Exchanging the bath solution with a hyposmotic (225 or 150 mosM) solution resulted in cellular swelling and selective activation of the K current (126 +/- 86 and 273 +/- 184% increase, respectively). Hyperosmotic solution changes (380 mosM) resulted in cell shrinkage and deactivation of the K current (44.2 +/- 21% decrease). Similar increases in the cell volume and whole cell current were observed on increasing (in perforated patch experiments) the chloride ion concentration (50 mM) in the pipette intracellular solution (127 +/- 63% increase). These changes were accompanied by marked shifts in the resting membrane voltage. We conclude that the K channels in these cells can respond to alteration in cellular osmolality or volume, resulting in changes in the whole cell current and resting voltage.

Voltage dependence of the Ca2+-activated K+channel KCa3.1 in human erythroleukemia cells

2013 ◽  
Vol 304 (9) ◽  
pp. C858-C872 ◽  
Author(s):  
Colin J. Stoneking ◽  
Oshini Shivakumar ◽  
David Nicholson Thomas ◽  
William H. Colledge ◽  
Michael J. Mason
Keyword(s):  
Voltage Dependence ◽  
Single Channel ◽  
K Channel ◽  
Whole Cell ◽  
Whole Cell Patch Clamp ◽  
Erythroleukemia Cells ◽  
Voltage Dependent ◽  
Hel Cells ◽  
Cell Current ◽  
Whole Cell Current

We have isolated a K+-selective, Ca2+-dependent whole cell current and single-channel correlate in the human erythroleukemia (HEL) cell line. The whole cell current was inhibited by the intermediate-conductance KCa3.1 inhibitors clotrimazole, TRAM-34, and charybdotoxin, unaffected by the small-conductance KCa2 family inhibitor apamin and the large-conductance KCa1.1 inhibitors paxilline and iberiotoxin, and augmented by NS309. The single-channel correlate of the whole cell current was blocked by TRAM-34 and clotrimazole, insensitive to paxilline, and augmented by NS309 and had a single-channel conductance in physiological K+gradients of ∼9 pS. RT-PCR revealed that the KCa3.1 gene, but not the KCa1.1 gene, was expressed in HEL cells. The KCa3.1 current, isolated in HEL cells under whole cell patch-clamp conditions, displayed an activated current component during depolarizing voltage steps from hyperpolarized holding potentials and tail currents upon repolarization, consistent with voltage-dependent modulation. This activated current increased with increasing voltage steps above −40 mV and was sensitive to inhibition by clotrimazole, TRAM-34, and charybdotoxin and insensitive to apamin, paxilline, and iberiotoxin. In single-channel experiments, depolarization resulted in an increase in open channel probability ( Po) of KCa3.1, with no increase in channel number. The voltage modulation of Powas an increasing monotonic function of voltage. In the absence of elevated Ca2+, voltage was ineffective at inducing channel activity in whole cell and single-channel experiments. These data indicate that KCa3.1 in HEL cells displays a unique form of voltage dependence modulating Po.

scholarly journals Effective Activation of BKCa Channels by QO-40 (5-(Chloromethyl)-3-(Naphthalen-1-yl)-2-(Trifluoromethyl)Pyrazolo [1,5-a]pyrimidin-7(4H)-one), Known to Be an Opener of KCNQ2/Q3 Channels

2021 ◽  
Vol 14 (5) ◽  
pp. 388
Author(s):  
Wei-Ting Chang ◽  
Sheng-Nan Wu
Keyword(s):  
Single Channel ◽  
Slow Component ◽  
Ionic Channel ◽  
Gh3 Cells ◽  
Bkca Channel ◽  
Bkca Channels ◽  
Gating Charge ◽  
Ramp Voltage ◽  
K Current ◽  
The Mean

QO-40 (5-(chloromethyl)-3-(naphthalene-1-yl)-2-(trifluoromethyl) pyrazolo[1,5-a]pyrimidin-7(4H)-one) is a novel and selective activator of KCNQ2/KCNQ3 K+ channels. However, it remains largely unknown whether this compound can modify any other type of plasmalemmal ionic channel. The effects of QO-40 on ion channels in pituitary GH3 lactotrophs were investigated in this study. QO-40 stimulated Ca2+-activated K+ current (IK(Ca)) with an EC50 value of 2.3 μM in these cells. QO-40-stimulated IK(Ca) was attenuated by the further addition of GAL-021 or paxilline but not by linopirdine or TRAM-34. In inside-out mode, this compound added to the intracellular leaflet of the detached patches stimulated large-conductance Ca2+-activated K+ (BKCa) channels with no change in single-channel conductance; however, there was a decrease in the slow component of the mean closed time of BKCa channels. The KD value required for the QO-40-mediated decrease in the slow component at the mean closure time was 1.96 μM. This compound shifted the steady-state activation curve of BKCa channels to a less positive voltage and decreased the gating charge of the channel. The application of QO-40 also increased the hysteretic strength of BKCa channels elicited by a long-lasting isosceles-triangular ramp voltage. In HEK293T cells expressing α-hSlo, QO-40 stimulated BKCa channel activity. Overall, these findings demonstrate that QO-40 can interact directly with the BKCa channel to increase the amplitude of IK(Ca) in GH3 cells.

scholarly journals Characterization of Inhibitory Effectiveness in Hyperpolarization-Activated Cation Currents by a Group of ent-Kaurane-Type Diterpenoids from Croton tonkinensis

2020 ◽  
Vol 21 (4) ◽  
pp. 1268 ◽  
Author(s):  
Ping-Chung Kuo ◽  
Yen-Chin Liu ◽  
Yi-Ching Lo ◽  
Sheng-Nan Wu
Keyword(s):  
Ionic Currents ◽  
Neuroendocrine Cells ◽  
Flowering Plant ◽  
Gh3 Cells ◽  
Functional Activities ◽  
Cation Current ◽  
Voltage Dependent ◽  
K Current ◽  
The Common

Croton is an extensive flowering plant genus in the spurge family, Euphorbiaceae. Three croton compounds with the common ent-kaurane skeleton have been purified from Croton tonkinensis. Methods: We examined any modifications of croton components (i.e., croton-01 [ent-18-acetoxy-7α-hydroxykaur-16-en-15-one], croton-02 [ent-7α,14β-dihydroxykaur-16-en-15-one] and croton-03 [ent-1β-acetoxy-7α,14β-dihydroxykaur-16-en-15-one] on either hyperpolarization-activated cation current (Ih) or erg-mediated K+ current identified in pituitary tumor (GH3) cells and in rat insulin-secreting (INS-1) cells via patch-clamp methods. Results: Addition of croton-01, croton-02, or croton-03 effectively and differentially depressed Ih amplitude. Croton-03 (3 μM) shifted the activation curve of Ih to a more negative potential by approximately 11 mV. The voltage-dependent hysteresis of Ih was also diminished by croton-03 administration. Croton-03-induced depression of Ih could not be attenuated by SQ-22536 (10 μM), an inhibitor of adenylate cyclase, but indeed reversed by oxaliplatin (10 μM). The Ih in INS-1 cells was also depressed effectively by croton-03. Conclusion: Our study highlights the evidence that these ent-kaurane diterpenoids might conceivably perturb these ionic currents through which they have high influence on the functional activities of endocrine or neuroendocrine cells.

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