Inwardly rectifying K+ channels in esophageal smooth muscle

2000 ◽  
Vol 279 (5) ◽  
pp. G951-G960 ◽  
Author(s):  
Junzhi Ji ◽  
Anne Marie F. Salapatek ◽  
Nicholas E. Diamant
Keyword(s):  
Membrane Potential ◽  
Longitudinal Muscle ◽  
Muscle Cells ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Patch Clamp Technique ◽  
Membrane Hyperpolarization ◽  
Inward Current ◽  
Inwardly Rectifying

The whole cell patch-clamp technique was used to investigate whether there were inwardly rectifying K+(Kir) channels in the longitudinal muscle of cat esophagus. Inward currents were observable on membrane hyperpolarization negative to the K+ equilibrium potential ( E k) in freshly isolated esophageal longitudinal muscle cells. The current-voltage relationship exhibited strong inward rectification with a reversal potential ( E rev) of −76.5 mV. Elevation of external K+ increased the inward current amplitude and positively shifted its E rev after the E k, suggesting that potassium ions carry this current. External Ba2+ and Cs+ inhibited this inward current, with hyperpolarization remarkably increasing the inhibition. The IC50 for Ba2+ and Cs+ at −60 mV was 2.9 and 1.6 mM, respectively. Furthermore, external Ba2+ of 10 μM moderately depolarized the resting membrane potential of the longitudinal muscle cells by 6.3 mV while inhibiting the inward rectification. We conclude that Kir channels are present in the longitudinal muscle of cat esophagus, where they contribute to its resting membrane potential.

Characterization of a Hyperpolarization-Activated Inward Current in Cajal-Retzius Cells in Rat Neonatal Neocortex

2000 ◽  
Vol 84 (3) ◽  
pp. 1681-1691 ◽  
Author(s):  
Werner Kilb ◽  
Heiko J. Luhmann
Keyword(s):  
Dissociation Constant ◽  
Reversal Potential ◽  
Hill Coefficient ◽  
Resting Membrane Potential ◽  
Patch Clamp Technique ◽  
Apparent Dissociation Constant ◽  
Membrane Hyperpolarization ◽  
Whole Cell ◽  
Inward Current ◽  
Retzius Cells

Cajal-Retzius cells are among the first neurons appearing during corticogenesis and play an important role in the establishment of cortical lamination. To characterize the hyperpolarization-activated inward current ( I h) and to investigate whether I h contributes to the relatively positive resting membrane potential (RMP) of these cells, we analyzed the properties of I h in visually identified Cajal-Retzius cells in cortical slices from neonatal rats using the whole cell patch-clamp technique. Membrane hyperpolarization to −90 mV activated a prominent inward current that was inhibited by 1 mM Cs+ and was insensitive to 1 mM Ba2+. The activation time constant for I h was strongly voltage dependent. In Na+-free solution, I h was reduced, indicating a contribution of Na+. An analysis of the tail currents revealed a reversal potential of −45.2 mV, corresponding to a permeability coefficient (pNa+/pK+) of 0.13. While an increase in the extracellular K+ concentration ([K+]e) enhances I h, it was reduced by a [K+]e decrease. This [K+]e dependence could not be explained by an effect on the electromotive force on K+ but suggested an additional extracellular binding site for K+ with an apparent dissociation constant of 7.2 mM. Complete Cl−substitution by Br−, I−, or NO3 − had no significant effect on I h, whereas a complete Cl−substitution by the organic compounds methylsulfate, isethionate, or gluconate reduced I h by ∼40%. The I h reduction observed in gluconate could be abolished by the addition of Cl−. The analysis of the [Cl−]e dependence of I h revealed a dissociation constant of 9.8 mM and a Hill-coefficient of 2.5, while the assumption of a gluconate-dependent I h reduction required an unreasonably high Hill-coefficient >20. An internal perfusion with the lidocaine derivative lidocaine N-ethyl bromide blocks I h within 1 min after establishment of the whole cell configuration. An inhibition of I h by 1 mM Cs+ was without an effect on RMP, action potential amplitude, threshold, width, or afterhyperpolarization. We conclude from these results that Cajal-Retzius cells express a prominent I hwith characteristic properties that does not contribute to the RMP.

Charybdotoxin block of Ca2+-activated K+ channels in colonic muscle depends on membrane potential dynamics

1998 ◽  
Vol 274 (3) ◽  
pp. C673-C680 ◽  
Author(s):  
Bret W. Frey ◽  
Andreas Carl ◽  
Nelson G. Publicover
Keyword(s):  
Membrane Potential ◽  
Longitudinal Muscle ◽  
Circular Muscle ◽  
Resting Membrane Potential ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Whole Cell Patch Clamp ◽  
Rate Of Increase ◽  
Voltage Dependent ◽  
Simple Kinetic Model

Charybdotoxin (ChTX) is a specific blocker of Ca2+-activated K+ channels. The voltage- and time-dependent dynamics of ChTX block were investigated using canine colonic myocytes and the whole cell patch-clamp technique with step and ramp depolarization protocols. During prolonged step depolarizations, K+ current slowly increased in the continued presence of ChTX (100 nM). The rate of increase depended on membrane potential with an e-fold change for every 60 mV. During ramp depolarizations, the effectiveness of ChTX block depended significantly on the rate of the ramp (50% at 0.01 V/s to 80% at 0.5 V/s). Results are consistent with a mechanism in which ChTX slowly “unbinds” in a voltage-dependent manner. A simple kinetic model was developed in which ChTX binds to both open and closed states. Slow unbinding is consistent with ChTX having little effect on electrical slow waves recorded from circular muscle while causing depolarization and contraction of longitudinal muscle, which displays more rapid “spikes.” Resting membrane potential and membrane potential dynamics are important determinants of ChTX action.

Functional expression of a Kir2.1-like inwardly rectifying potassium channel in mouse mammary secretory cells

2014 ◽  
Vol 306 (3) ◽  
pp. C230-C240 ◽  
Author(s):  
Akihiro Kamikawa ◽  
Toru Ishikawa
Keyword(s):  
Mammary Gland ◽  
Ionic Composition ◽  
Functional Expression ◽  
Mouse Mammary Gland ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Pcr Analysis ◽  
Secretory Cells ◽  
Patch Clamp Technique ◽  
Inwardly Rectifying

K+ channels in mammary secretory (MS) cells are believed to play a role in transcellular electrolyte transport and thus determining ionic composition of the aqueous phase of milk. However, direct evidence for specific K+ channel activity in native MS cells is lacking at the single-cell level. Here, we show for the first time that an inwardly rectifying K+ (Kir) channel is functionally expressed in fully differentiated MS cells that were freshly isolated from the mammary gland of lactating mice. Using the standard whole cell patch-clamp technique, we found that mouse MS cells consistently displayed a K+ current, whose electrophysiological properties are similar to those previously reported for Kir2.x channels, particularly Kir2.1: 1) current-voltage relationship with strong inward rectification, 2) slope conductance approximately proportional to the square root of external K+ concentration, 3) voltage- and time-dependent and high-affinity block by external Ba2+, and 4) voltage-dependent inhibition by external Cs+. Accordingly, RT-PCR analysis revealed the gene expression of Kir2.1, but not Kir2.2, Kir2.3, and Kir2.4, in lactating mouse mammary gland, and immunohistochemical staining showed Kir2.1 protein expression in the secretory cells. Cell-attached patch recordings from MS cells revealed that a 31-pS K+ channel with strong inward rectification was likely active at the resting membrane potential. Collectively, the present work demonstrates that a functional Kir2.1-like channel is expressed in lactating mouse MS cells. We propose that the channel might be involved, at least in part, in secretion and/or preservation of ionic components of milk stored into the lumen of these cells.

Demonstration of an electrogenic Na+-K+ pump in mouse spleen macrophages

1983 ◽  
Vol 245 (3) ◽  
pp. C184-C188 ◽  
Author(s):  
E. K. Gallin ◽  
D. R. Livengood
Keyword(s):  
Membrane Potential ◽  
Mouse Spleen ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Free Medium ◽  
Membrane Hyperpolarization ◽  
Induced Membrane ◽  
Current Voltage ◽  
Pump Activity ◽  
Subsequent Exposure

The effect of ouabain on the membrane potential of cultured mouse spleen macrophages was examined. Ouabain (10(-3) M) induced a membrane depolarization (6.6 mV) in 18 of 19 cells studied that occurred within several minutes after exposure and was not associated with significant changes in current-voltage relationships. In related studies, cells were placed in K+-free medium in the cold for 2 h to block pump activity. Subsequent exposure to K+ at 37 degrees C resulted in a membrane hyperpolarization. However, addition of K+ also enhanced inward rectification. To differentiate between the effect of K+ on the Na+-K+ pump and its action on inward rectification, two types of experiments were done. Studies performed in the presence of barium, which blocks inward rectification, demonstrated a K+-induced hyperpolarization with no changes in rectification. Additional studies examined the effects of rubidium on Na+-loaded cells. Rubidium, which blocks inward rectification but substitutes for K+ in activating the Na+-K+ pump, induced membrane hyperpolarizations that were reversed by addition of ouabain. These data indicate that macrophages exhibit an electrogenic Na+-K+ pump, which probably contributes to the resting membrane potential under steady-state conditions and can be activated under conditions designed to Na+ load the cells. In addition, they demonstrate that increasing extracellular K+ enhances inward rectification in macrophages.

Identification of a spontaneously active, Na+-permeable channel in guinea pig gallbladder smooth muscle

2005 ◽  
Vol 289 (3) ◽  
pp. G501-G507 ◽  
Author(s):  
Georgi V. Petkov ◽  
Onesmo B. Balemba ◽  
Mark T. Nelson ◽  
Gary M. Mawe
Keyword(s):  
Smooth Muscle ◽  
Membrane Potential ◽  
Guinea Pig ◽  
Action Potential ◽  
Equilibrium Potential ◽  
Resting Membrane Potential ◽  
Action Potential Generation ◽  
Potential Generation ◽  
Membrane Hyperpolarization ◽  
Permeable Channel

The action potential in gallbladder smooth muscle (GBSM) is caused by Ca2+ entry through voltage-dependent Ca2+ channels (VDCC), which contributes to the GBSM contractions. Action potential generation in GBSM is critically dependent on the resting membrane potential (about −50 mV), which is ∼35 mV more positive of the K+ equilibrium potential. We hypothesized that a tonic, depolarizing conductance is present in GBSM and contributes to the regulation of the resting membrane potential and action potential frequency. GBSM cells were isolated from guinea pig gallbladders, and the whole cell patch-camp technique was used to record membrane currents. After eliminating the contribution of VDCC and K+ channels, we identified a novel spontaneously active cation conductance ( Icat) in GBSM. This Icat was mediated predominantly by influx of Na+. Na+ substitution with N-methyl-d-glucamine (NMDG), a large relatively impermeant cation, caused a negative shift in the reversal potential of the ramp current and reduced the amplitude of the inward current at −50 mV by 65%. Membrane potential recordings with intracellular microelectrodes or in current-clamp mode of the patch-clamp technique indicated that the inhibition of Icat conductance by NMDG is associated with membrane hyperpolarization and inhibition of action potentials. Extracellular Ca2+, Mg2+, and Gd3+ attenuated the Icat in GBSM. Muscarinic stimulation did not activate the Icat. Our results indicate that, in GBSM, an Na+-permeable channel contributes to the maintenance of the resting membrane potential and action potential generation and therefore plays a critical role in the regulation of GBSM excitability and contractility.

Electrophysiological response of rat ventricular myocytes to acidosis

2002 ◽  
Vol 283 (1) ◽  
pp. H412-H422 ◽  
Author(s):  
Kimiaki Komukai ◽  
Fabien Brette ◽  
Caroline Pascarel ◽  
Clive H. Orchard
Keyword(s):  
Membrane Potential ◽  
Action Potential ◽  
Ventricular Myocytes ◽  
Resting Potential ◽  
Disulfonic Acid ◽  
Resting Membrane Potential ◽  
Delayed Rectifier ◽  
Patch Clamp Technique ◽  
Electrophysiological Response ◽  
Inwardly Rectifying

The effects of acidosis on the action potential, resting potential, L-type Ca2+( I Ca), inward rectifier potassium ( I K1), delayed rectifier potassium ( I K), steady-state ( I SS), and inwardly rectifying chloride ( I Cl,ir) currents of rat subepicardial (Epi) and subendocardial (Endo) ventricular myocytes were investigated using the patch-clamp technique. Action potential duration was shorter in Epi than in Endo cells. Acidosis (extracellular pH decreased from 7.4 to 6.5) depolarized the resting membrane potential and prolonged the time for 50% repolarization of the action potential in Epi and Endo cells, although the prolongation was larger in Endo cells. At control pH, I Ca, I K1, and I SS were not significantly different in Epi and Endo cells, but I K was larger in Epi cells. Acidosis did not alter I Ca, I K1, or I K but decreased I SS; this decrease was larger in Endo cells. It is suggested that the acidosis-induced decrease in I SS underlies the prolongation of the action potential. I Cl,ir at control pH was Cd2+ sensitive but 4,4′-disothiocyanato-stilbene-2,2′-disulfonic acid resistant. Acidosis increased I Cl,ir; it is suggested that the acidosis-induced increase in I Cl,ir underlies the depolarization of the resting membrane potential.

Volume-activated chloride currents in interstitial cells of Cajal

2005 ◽  
Vol 289 (5) ◽  
pp. G791-G797 ◽  
Author(s):  
Sung Jin Park ◽  
Catherine M. Mckay ◽  
Yaohui Zhu ◽  
Jan D. Huizinga
Keyword(s):  
Membrane Potential ◽  
Slow Wave ◽  
Interstitial Cells Of Cajal ◽  
Morphological Changes ◽  
Interstitial Cells ◽  
Equilibrium Potential ◽  
Resting Membrane Potential ◽  
Hypotonic Solution ◽  
Patch Clamp Technique ◽  
Cells Of Cajal

Interstitial cells of Cajal (ICC) undergo marked morphological changes on contraction of the musculature, making it essential to understand properties of mechanosensitive ion channels. The whole cell patch-clamp technique was used to identify and to characterize volume-activated Cl− currents in ICC cultured through the explant technique. Hypotonic solutions (≈210 mosM) activated an outwardly rectifying current, which reversed near the equilibrium potential for Cl−. Time-dependent inactivation occurred only at pulse potentials of +80 mV, with a time constant of 478 ± 182 ms. The degree of outward rectification was calculated using a rectification index, the ratio between the slope conductances of +65 and −55 mV, which was 13.9 ± 1.5 at 76 mM initial extracellular Cl− concentration. The sequence of relative anion permeability of the outwardly rectifying Cl− channel was I− > Cl− > aspartate−. The chloride channel blockers, DIDS and 5-nitro-2-(3-phenlypropl-amino)benzoic acid, caused a voltage-dependent block of the outwardly rectifying Cl− current, inhibition occurring primarily at depolarized potentials. On exposure to hypotonic solution, the slope conductance significantly increased at the resting membrane potential (−70 mV) from 1.2 ± 0.2 to 2.0 ± 0.4 nS and at the slow-wave plateau potential (−35 mV) from 2.1 ± 0.3 to 5.0 ± 1.0 nS. The current was constitutively active in ICC and contributed to the resting membrane potential and excitability at the slow-wave plateau. In conclusion, swelling or volume change will depolarize ICC through activation of outwardly rectifying chloride channels, thereby increasing cell excitability.

Screening Technologies for Inward Rectifier Potassium Channels: Discovery of New Blockers and Activators

2020 ◽  
Vol 25 (5) ◽  
pp. 420-433 ◽  
Author(s):  
Kenneth B. Walsh
Keyword(s):  
Membrane Potential ◽  
G Protein ◽  
Critical Role ◽  
Inward Rectifier ◽  
Resting Potential ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Kir Channels ◽  
Inward Rectifier Potassium Channels

K+ channels play a critical role in maintaining the normal electrical activity of excitable cells by setting the cell resting membrane potential and by determining the shape and duration of the action potential. In nonexcitable cells, K+ channels establish electrochemical gradients necessary for maintaining salt and volume homeostasis of body fluids. Inward rectifier K+ (Kir) channels typically conduct larger inward currents than outward currents, resulting in an inwardly rectifying current versus voltage relationship. This property of inward rectification results from the voltage-dependent block of the channels by intracellular polyvalent cations and makes these channels uniquely designed for maintaining the resting potential near the K+ equilibrium potential (EK). The Kir family of channels consist of seven subfamilies of channels (Kir1.x through Kir7.x) that include the classic inward rectifier (Kir2.x) channel, the G-protein-gated inward rectifier K+ (GIRK) (Kir3.x), and the adenosine triphosphate (ATP)-sensitive (KATP) (Kir 6.x) channels as well as the renal Kir1.1 (ROMK), Kir4.1, and Kir7.1 channels. These channels not only function to regulate electrical/electrolyte transport activity, but also serve as effector molecules for G-protein-coupled receptors (GPCRs) and as molecular sensors for cell metabolism. Of significance, Kir channels represent promising pharmacological targets for treating a number of clinical conditions, including cardiac arrhythmias, anxiety, chronic pain, and hypertension. This review provides a brief background on the structure, function, and pharmacology of Kir channels and then focuses on describing and evaluating current high-throughput screening (HTS) technologies, such as membrane potential-sensitive fluorescent dye assays, ion flux measurements, and automated patch clamp systems used for Kir channel drug discovery.

scholarly journals Ca2+-activated K+ Channels in Murine Endothelial Cells: Block by Intracellular Calcium and Magnesium

2008 ◽  
Vol 131 (2) ◽  
pp. 125-135 ◽  
Author(s):  
Jonathan Ledoux ◽  
Adrian D. Bonev ◽  
Mark T. Nelson
Keyword(s):  
Endothelial Cells ◽  
Membrane Potential ◽  
Intracellular Calcium ◽  
Membrane Current ◽  
Mesenteric Arteries ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Resting Membrane Potential ◽  
Additional Increase ◽  
K Channels

The intermediate (IKCa) and small (SKCa) conductance Ca2+-sensitive K+ channels in endothelial cells (ECs) modulate vascular diameter through regulation of EC membrane potential. However, contribution of IKCa and SKCa channels to membrane current and potential in native endothelial cells remains unclear. In freshly isolated endothelial cells from mouse aorta dialyzed with 3 μM free [Ca2+]i and 1 mM free [Mg2+]i, membrane currents reversed at the potassium equilibrium potential and exhibited an inward rectification at positive membrane potentials. Blockers of large-conductance, Ca2+-sensitive potassium (BKCa) and strong inward rectifier potassium (Kir) channels did not affect the membrane current. However, blockers of IKCa channels, charybdotoxin (ChTX), and of SKCa channels, apamin (Ap), significantly reduced the whole-cell current. Although IKCa and SKCa channels are intrinsically voltage independent, ChTX- and Ap-sensitive currents decreased steeply with membrane potential depolarization. Removal of intracellular Mg2+ significantly increased these currents. Moreover, concomitant reduction of the [Ca2+]i to 1 μM caused an additional increase in ChTX- and Ap-sensitive currents so that the currents exhibited theoretical outward rectification. Block of IKCa and SKCa channels caused a significant endothelial membrane potential depolarization (≈11 mV) and decrease in [Ca2+]i in mesenteric arteries in the absence of an agonist. These results indicate that [Ca2+]i can both activate and block IKCa and SKCa channels in endothelial cells, and that these channels regulate the resting membrane potential and intracellular calcium in native endothelium.

Descending vasa recta endothelia express inward rectifier potassium channels

2007 ◽  
Vol 293 (4) ◽  
pp. F1248-F1255 ◽  
Author(s):  
Chunhua Cao ◽  
Whaseon Lee-Kwon ◽  
Kristie Payne ◽  
Aurélie Edwards ◽  
Thomas L. Pallone
Keyword(s):  
Endothelial Cells ◽  
Membrane Potential ◽  
Potassium Channels ◽  
Inward Rectifier ◽  
Equilibrium Potential ◽  
Dependent Manner ◽  
Membrane Hyperpolarization ◽  
Inward Rectifier Potassium Channels ◽  
Vasa Recta ◽  
Inwardly Rectifying

Descending vasa recta (DVR) are capillary-sized microvessels that supply blood flow to the renal medulla. They are composed of contractile pericytes and endothelial cells. In this study, we used the whole cell patch-clamp method to determine whether inward rectifier potassium channels (KIR) exist in the endothelia, affect membrane potential, and modulate intracellular Ca2+ concentration ([Ca2+]cyt). The endothelium was accessed for electrophysiology by removing abluminal pericytes from collagenase-digested vessels. KIR currents were recorded using symmetrical 140 mM K+ solutions that served to maximize currents and eliminate cell-to-cell coupling by closing gap junctions. Large, inwardly rectifying currents were observed at membrane potentials below the equilibrium potential for K+. Ba2+ potently inhibited those currents in a voltage-dependent manner, with affinity k = 0.18, 0.33, 0.60, and 1.20 μM at −160, −120, −80, and −40 mV, respectively. Cs+ also blocked those currents with k = 20, 48, 253, and 1,856 μM at −160, −120, −80, and −40 mV, respectively. In the presence of 1 mM ouabain, increasing extracellular K+ concentration from 5 to 10 mM hyperpolarized endothelial membrane potential by 15 mV and raised endothelial [Ca2+]cyt. Both the K+-induced membrane hyperpolarization and the [Ca2+]cyt elevation were reversed by Ba2+. Immunochemical staining verified that both pericytes and endothelial cells of DVR express KIR2.1, KIR2.2, and KIR2.3 subunits. We conclude that strong, inwardly rectifying KIR2.x isoforms are expressed in DVR and mediate K+-induced hyperpolarization of the endothelium.

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