Activation of whole cell currents in isolated human jejunal circular smooth muscle cells by carbon monoxide

1993 ◽  
Vol 264 (6) ◽  
pp. G1184-G1189 ◽  
Author(s):  
G. Farrugia ◽  
W. A. Irons ◽  
J. L. Rae ◽  
M. G. Sarr ◽  
J. H. Szurszewski
Keyword(s):  
Carbon Monoxide ◽  
Smooth Muscle ◽  
Membrane Potential ◽  
Muscle Cells ◽  
Patch Clamp Technique ◽  
Current Increase ◽  
Membrane Hyperpolarization ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

Carbon monoxide (CO) is a low molecular weight oxide produced endogenously from fatty acids and heme protein. A physiological role for CO has been suggested for vascular smooth muscle, hemostasis, and olfactory neurons, but direct evidence is lacking. Heme oxygenase, which catalyzes the formation of CO from heme proteins, is present in small intestinal smooth muscle. The effect of 1% CO on whole cell currents in normal human jejunal circular muscle cells was studied with the use of a perforated patch-clamp technique. A 1% CO-containing Krebs solution caused an initial and transient increase in whole cell current in 20 of 22 cells tested (175 +/- 40%, mean +/- SE) and a transient hyperpolarization (15.6 +/- 3.6 mV, mean +/- SE) of the membrane potential. During prolonged recordings, 1% CO evoked ongoing cyclic increases and decreases in the whole cell current. Each current increase was accompanied by a sharp membrane hyperpolarization. These data suggest that CO may modulate whole cell potassium current and membrane potential.

Whole cell current and membrane potential regulation by a human smooth muscle mechanosensitive calcium channel

2000 ◽  
Vol 279 (6) ◽  
pp. G1155-G1161 ◽  
Author(s):  
Adrian N. Holm ◽  
Adam Rich ◽  
Michael G. Sarr ◽  
Gianrico Farrugia
Keyword(s):  
Smooth Muscle ◽  
Membrane Potential ◽  
Smooth Muscle Cells ◽  
Outward Current ◽  
Muscle Cells ◽  
Membrane Hyperpolarization ◽  
Whole Cell ◽  
Circular Smooth Muscle ◽  
Channel Current ◽  
Cell Current

Mechanotransduction is required for a wide variety of biological functions. The aim of this study was to determine the effect of activation of a mechanosensitive Ca2+ channel, present in human jejunal circular smooth muscle cells, on whole cell currents and on membrane potential. Currents were recorded using patch-clamp techniques, and perfusion of the bath (10 ml/min, 30 s) was used to mechanoactivate the L-type Ca2+ channel. Perfusion resulted in activation of L-type Ca2+ channels and an increase in outward current from 664 ± 57 to 773 ± 72 pA at +60 mV. Membrane potential hyperpolarized from −42 ± 4 to −50 ± 5 mV. In the presence of nifedipine (10 μM), there was no increase in outward current or change in membrane potential with perfusion. In the presence of charybdotoxin or iberiotoxin, perfusion of the bath did not increase outward current or change membrane potential. A model is proposed in which mechanoactivation of an L-type Ca2+ channel current in human jejunal circular smooth muscle cells results in increased Ca2+ entry and cell contraction. Ca2+ entry activates large-conductance Ca2+-activated K+channels, resulting in membrane hyperpolarization and relaxation.

Distribution of heme oxygenase and effects of exogenous carbon monoxide in canine jejunum

1998 ◽  
Vol 274 (2) ◽  
pp. G350-G358 ◽  
Author(s):  
G. Farrugia ◽  
S. M. Miller ◽  
A. Rich ◽  
X. Liu ◽  
M. D. Maines ◽  
...  
Keyword(s):  
Carbon Monoxide ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Heme Oxygenase ◽  
Neuronal Cell ◽  
Muscle Cells ◽  
Nerve Fibers ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

Carbon monoxide (CO) has been postulated to be a messenger in the gastrointestinal tract. The aims of this study were to determine the distribution of heme oxygenase (HO), the source for endogenous CO in the canine jejunum, and to determine the effects of CO on jejunal circular smooth muscle cells. HO-2 isoform was present in a population of myenteric and submucosal neuronal cell bodies, in nerve fibers innervating the muscle layers, and in smooth muscle cells. HO-1 isozyme was not detected in the canine jejunum. Exogenous CO increased whole cell current by 285 ± 86%, hyperpolarized the membrane potential by 8.5 ± 2.9 mV, and increased guanosine 3′,5′-cyclic monophosphate (cGMP) levels in smooth muscle cells. 8-Bromo- cGMP also increased the whole cell current. The data suggest that endogenous activity of HO-2 may be a source of CO in the canine jejunum and that exogenously applied CO can modulate intestinal smooth muscle electrical activity. It is therefore reasonable to suggest a role for endogenously produced CO as a messenger in the canine jejunum.

scholarly journals [Ca2+]i oscillations in human sperm are triggered in the flagellum by membrane potential-sensitive activity of CatSper

2020 ◽  
Author(s):  
Elis Torrezan-Nitao ◽  
Sean G Brown ◽  
Esperanza Mata-Martínez ◽  
Claudia L Treviño ◽  
Christopher Barratt ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Large Scale ◽  
Conflicts Of Interest ◽  
Human Sperm ◽  
Equilibrium Potential ◽  
Current Clamp ◽  
Whole Cell ◽  
Cell Current ◽  
Whole Cell Current

Abstract STUDY QUESTION How are progesterone (P4)-induced repetitive intracellular Ca2+ concentration ([Ca2+]i) signals (oscillations) in human sperm generated? SUMMARY ANSWER P4-induced [Ca2+]i oscillations are generated in the flagellum by membrane potential (Vm)-sensitive Ca2+-influx through CatSper channels. WHAT IS KNOWN ALREADY A subset of human sperm display [Ca2+]i oscillations that regulate flagellar beating and acrosome reaction. Although pharmacological manipulations indicate involvement of stored Ca2+ in these oscillations, influx of extracellular Ca2+ is also required. STUDY DESIGN, SIZE, DURATION This was a laboratory study that used >20 sperm donors and involved more than 100 separate experiments and analysis of more than 1000 individual cells over a period of 2 years. PARTICIPANTS/MATERIALS, SETTING, METHODS Semen donors and patients were recruited in accordance with local ethics approval from Birmingham University and Tayside ethics committees. [Ca2+]i responses and Vm of individual cells were examined by fluorescence imaging and whole-cell current clamp. MAIN RESULTS AND THE ROLE OF CHANCE P4-induced [Ca2+]i oscillations originated in the flagellum, spreading to the neck and head (latency of 1–2 s). K+-ionophore valinomycin (1 µM) was used to investigate the role of membrane potential (Vm). Direct assessment by whole-cell current-clamp confirmed that Vm in valinomycin-exposed cells was determined primarily by K+ equilibrium potential (EK) and was rapidly ‘reset’ upon manipulation of [K+]o. Pre-treatment of sperm with valinomycin ([K+]o = 5.4 mM) had no effect on the P4-induced [Ca2+] transient (P = 0.95; eight experiments), but application of valinomycin to P4-pretreated sperm suppressed activity in 82% of oscillating cells (n = 257; P = 5 × 10−55 compared to control) and significantly reduced both the amplitude and frequency of persisting oscillations (P = 0.0001). Upon valinomycin washout, oscillations re-started in most cells. When valinomycin was applied in saline with elevated [K+], the inhibitory effect of valinomycin was reduced and was dependent on EK (P = 10−25). Amplitude and frequency of [Ca2+]i oscillations that persisted in the presence of valinomycin showed similar sensitivity to EK (P < 0.01). The CatSper inhibitor RU1968 (4.8 and 11 µM) caused immediate and reversible arrest of activity in 36% and 96% of oscillating cells, respectively (P < 10−10). Quinidine (300 µM) which blocks the sperm K+ current (IKsper) completely, inhibited [Ca2+]i oscillations. LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION This was an in-vitro study and caution must be taken when extrapolating these results to in-vivo regulation of sperm. WIDER IMPLICATIONS OF THE FINDINGS [Ca2+]i oscillations in human sperm are functionally important and their absence is associated with failed fertilisation at IVF. The data reported here provide new understanding of the mechanisms that underlie the regulation and generation (or failure) of these oscillations. STUDY FUNDING/COMPETING INTEREST(S) E.T.-N. was in receipt of a postgraduate scholarship from the CAPES Foundation (Ministry of Education, Brazil). E.M-M received travel funds from the Programa de Apoyo a los Estudios de Posgrado (Maestria y Doctorado en Ciencias Bioquimicas-Universidad Autonoma de Mexico). SGB and CLRB are recipients of a Chief Scientist Office (NHS Scotland) grant TCS/17/28. The authors have no conflicts of interest.

Chronic carbon monoxide enhanced IbTx-sensitive currents in rat resistance pulmonary artery smooth muscle cells

2002 ◽  
Vol 283 (1) ◽  
pp. L120-L129 ◽  
Author(s):  
Eric Dubuis ◽  
Mathieu Gautier ◽  
Alexandre Melin ◽  
Manuel Rebocho ◽  
Catherine Girardin ◽  
...  
Keyword(s):  
Carbon Monoxide ◽  
Smooth Muscle ◽  
Membrane Potential ◽  
Pulmonary Artery ◽  
Smooth Muscle Cells ◽  
Outward Current ◽  
Muscle Cells ◽  
Membrane Resistance ◽  
Resting Membrane Potential ◽  
Whole Cell Patch Clamp

Exogenous carbon monoxide (CO) can induce pulmonary vasodilation by acting directly on pulmonary artery (PA) smooth muscle cells. We investigated the contribution of K+ channels to the regulation of resistance PA resting membrane potential on control (PAC) rats and rats exposed to CO for 3 wk at 530 parts/million, labeled as PACO rats. Whole cell patch-clamp experiments revealed that the resting membrane potential of PACO cells was more negative than that of PAC cells. This was associated with a decrease of membrane resistance in PACO cells. Additional analysis showed that outward current density in PACO cells was higher (50% at +60 mV) than in PAC cells. This was linked to an increase of iberiotoxin (IbTx)-sensitive current. Chronic CO hyperpolarized membrane of pressurized PA from −46.9 ± 1.2 to −56.4 ± 2.6 mV. Additionally, IbTx significantly depolarized membrane of smooth muscle cells from PACO arteries but not from PAC arteries. The present study provides initial evidence of an increase of Ca2+-activated K+ current in smooth muscle cells from PA of rats exposed to chronic CO.

Electrophysiological effects of O2−· on the plasma membrane in vascular endothelial cells

2005 ◽  
Vol 289 (6) ◽  
pp. H2379-H2386 ◽  
Author(s):  
Anna K. Brzezinska ◽  
Nicole Lohr ◽  
William M. Chilian
Keyword(s):  
Endothelial Cells ◽  
Ion Channels ◽  
Vascular Endothelial Cells ◽  
Vascular Dysfunction ◽  
Vascular Endothelial ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Cell Current ◽  
Electrophysiological Effects ◽  
Whole Cell Current

Vascular dysfunction is a hallmark of many diseases, including coronary heart disease, stroke, and diabetes. The underlying mechanisms of these disorders are intimately associated with an increase in oxidative stress and excess generation of reactive oxygen species. Here, we report that the anionic free radical, superoxide (O2−·), directly affects the function of ion channels in vascular endothelial cells. Vascular endothelial cells were exposed to O2−· under physiological, symmetrical chloride and chloride-free conditions. Superoxide was generated from the reaction of xanthine (0.2 mM) and xanthine oxidase (0.1, 1, and 10 mU/ml) while its effects were determined with the whole cell mode of the patch-clamp technique. Inhibitors of K+ and Cl− channels were used to determine the role of these ion channels in mediating the electrophysiological effects of superoxide. The addition of O2−· caused a dose-dependent depolarization of endothelial cells and activation of the whole cell current. Activation of superoxide-dependent current was observed in the presence of inhibitors of K+ channels, Ba2+ (100 μM) or iberiotoxin (100 nM), and was not affected by inhibitors of nonselective cation channels, La3+, or by inhibition of the Cl−/HCO3− transporter by bumetanide. The inhibitors of the Cl− channel, NPPB (0.1 mM) or DIDS (100 μM), partially prevented activation of superoxide-dependent current but were unable to reverse it. The effects of superoxide on the amplitude of whole cell current were prevented and reversed by superoxide dismutase. Taken together, these results suggest that superoxide directly affects the function of ion channels in vascular endothelium but the mechanisms of its modulatory effects remain unresolved.

scholarly journals Modulation of calcium channels in arterial smooth muscle cells by dihydropyridine enantiomers.

1993 ◽  
Vol 101 (3) ◽  
pp. 393-410 ◽  
Author(s):  
S Hering ◽  
A D Hughes ◽  
E N Timin ◽  
T B Bolton
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Pulse Train ◽  
Muscle Cells ◽  
Drug Action ◽  
Bay K 8644 ◽  
Patch Clamp Technique ◽  
Current Increase ◽  
Inactivation Curve ◽  
Mean Time

The actions of the optical enantiomers of BAY K 8644 and Sandoz 202,791 were studied on barium inward currents recorded using the whole-cell configuration of the patch clamp technique from enzymatically isolated smooth muscle cells from the rabbit ear artery. The enantiomers were applied by bath perfusion or rapidly by a concentration jump technique, which enabled the study of drug action under equilibrium and nonequilibrium conditions. A larger effect of agonists was seen on peak inward current in 110 mM Ba when small rather than large depolarizations were applied. The midpoint voltage of the steady-state inactivation curve of IBa was -12.8 +/- 1.9 mV (n = 4) in the absence of drug, -16.4 +/- 2.5 mV (n = 4) in 1 microM (+)202,791, and -31.4 +/- 0.4 mV (n = 4) in 1 microM (-)202,791. The rate of onset of action of the agonist and antagonist enantiomers of BAY K 8644 and Sandoz 202,791 was studied by rapid application during 20-ms depolarizing steps from different holding potentials to +30 mV at 1 or 0.2 Hz. The drugs were applied as concentration jumps between two single pulses of a pulse train. The rates of onset of drug action on peak IBa during a 1-Hz pulse train were concentration dependent over the range of 100 nM-3 microM for both (+) and (-)202,791. The rate of onset of inhibition of peak current by antagonist enantiomers was not significantly influenced by the test pulse frequency. At a holding potential of -60 mV, the onset rate of the increase in peak IBa on application of 1 microM of agonist enantiomers (+)202,791 or (-)BAY K 8644 during a train of pulses occurred with mean time constants of 2.1 +/- 0.7 s (n = 7) and 2.3 +/- 0.2 s (n = 4), respectively. The onset of current increase on application of 1 microM (+)202,791 during a single voltage clamp step to 20 mV was faster, with a mean time constant of 380 +/- 80 ms (n = 3).

PGE2 sensitizes cultured pulmonary vagal sensory neurons to chemical and electrical stimuli

2002 ◽  
Vol 93 (4) ◽  
pp. 1419-1428 ◽  
Author(s):  
Kevin Kwong ◽  
Lu-Yuan Lee
Keyword(s):  
Current Density ◽  
Sensory Nerves ◽  
In Vivo Studies ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Prostanoid Receptor ◽  
C Fibers ◽  
Mediators Of Inflammation ◽  
Cell Current ◽  
Whole Cell Current

Mediators of inflammation, such as PGE2, are known to sensitize the airways to inhaled irritants and circulating autacoids. Evidence from in vivo studies has shown the involvement of vagal pulmonary C-fiber afferents in the PGE2-elicited airway hypersensitivity. However, whether PGE2 acts directly on these sensory nerves is unclear. The present study aimed to investigate whether PGE2 has direct potentiating effects on nodose and jugular pulmonary C neurons cultured from adult Sprague-Dawley rats and, if so, determine whether the EP2 prostanoid receptor is involved. Pulmonary neurons were identified by retrograde labeling with a fluorescent tracer 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate. Using perforated patch-clamp technique, our results showed that 1) PGE2 pretreatment (1 μM) increased the whole cell current density elicited by capsaicin and phenylbiguanide, chemical agents known to stimulate pulmonary C fibers; 2) selective activation of the EP2 prostanoid receptor by butaprost (3–10 μM) increased the whole cell current density elicited by capsaicin; and 3) PGE2, as well as butaprost, increased the number of action potentials evoked by current injection. Therefore, we conclude that PGE2 directly sensitizes vagal pulmonary C neurons to chemical and electrical stimulation. Furthermore, butaprost modulates the neurons in a manner similar to that of PGE2, suggesting that the effects of PGE2 are mediated, at least in part, through the EP2 prostanoid receptor.

Nitric oxide modulates a calcium-activated potassium current in muscle cells from opossum esophagus

1995 ◽  
Vol 269 (4) ◽  
pp. G606-G612 ◽  
Author(s):  
J. A. Murray ◽  
E. F. Shibata ◽  
T. L. Buresh ◽  
H. Picken ◽  
B. W. O'Meara ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Protein Kinase ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Protein Kinase G ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Circular Smooth Muscle ◽  
K Current

Nitric oxide mediates nerve-induced hyperpolarization of circular smooth muscle of the esophagus. Two mechanisms are proposed to explain this hyperpolarization: an increase in K+ current or a decrease in Cl- current. These studies test the hypothesis that nitric oxide increases a K+ current in esophageal smooth muscle. Three outward K+ currents are present in circular smooth muscle cells from the opossum esophagus. One current is a Ca(2+)-activated K+ current (IKCa2+). This current is inhibited by charybdotoxin. Whole cell currents were recorded from isolated opossum esophageal smooth muscle cells using the whole cell patch-clamp technique. These studies showed that IKCa2+ is activated at potentials more positive than -30 mV. Bath application of S-nitroso-L-cysteine increased IKCa2+ by 50% above control levels throughout the entire activation range of potentials. The enhanced current was reversible on washout. Either charybdotoxin, an inhibitor of IKCa2+, or (R)-p-8-(4-chloropenylthio)-guanosine 3',5'-cyclic monophosphorothioate, an inhibitor of protein kinase G, antagonized the increase in outward current induced by S-nitroso-L-cysteine. These data suggest that nitric oxide activates IKCa2+ via the guanosine 3',5'-cyclic monophosphate-protein kinase G signal transduction pathway.

scholarly journals Extracellular pH and intracellular phosphatidylinositol 4,5-bisphosphate control Cl− currents in guinea pig detrusor smooth muscle cells

2019 ◽  
Vol 317 (6) ◽  
pp. C1268-C1277 ◽  
Author(s):  
Viktor Yarotskyy ◽  
John Malysz ◽  
Georgi V. Petkov
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Extracellular Ph ◽  
Complex Nature ◽  
Airway Smooth Muscle Cells ◽  
Detrusor Smooth Muscle ◽  
Patch Clamp Technique ◽  
Whole Cell

Cl− channels serve as key regulators of excitability and contractility in vascular, intestinal, and airway smooth muscle cells. We recently reported a Cl− conductance in detrusor smooth muscle (DSM) cells. Here, we used the whole cell patch-clamp technique to further characterize biophysical properties and physiological regulators of the Cl− current in freshly isolated guinea pig DSM cells. The Cl− current demonstrated outward rectification arising from voltage-dependent gating of Cl− channels rather than the Cl− transmembrane gradient. An exposure of DSM cells to hypotonic extracellular solution (Δ 165 mOsm challenge) did not increase the Cl− current providing strong evidence that volume-regulated anion channels do not contribute to the Cl− current in DSM cells. The Cl− current was monotonically dependent on extracellular pH, larger and lower in magnitude at acidic (5.0) and basic pH (8.5) values, respectively. Additionally, intracellularly applied phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] analog [PI(4,5)P2-diC8] increased the average Cl− current density by approximately threefold in a voltage-independent manner. The magnitude of the DSM whole cell Cl− current did not depend on the cell surface area (cell capacitance) regardless of the presence or absence of PI(4,5)P2-diC8, an intriguing finding that underscores the complex nature of Cl− channel expression and function in DSM cells. Removal of both extracellular Ca2+ and Mg2+ did not affect the DSM whole cell Cl− current, whereas Gd3+ (1 mM) potentiated the current. Collectively, our recent and present findings strongly suggest that Cl− channels are critical regulators of DSM excitability and are regulated by extracellular pH, Gd3+, and PI(4,5)P2.

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