scholarly journals Characterization of norepinephrine-evoked inward currents in interstitial cells isolated from the rabbit urethra

2002 ◽  
Vol 283 (3) ◽  
pp. C885-C894 ◽  
Author(s):  
G. P. Sergeant ◽  
K. D. Thornbury ◽  
N. G. McHale ◽  
M. A. Hollywood
Keyword(s):  
Reversal Potential ◽  
Cyclopiazonic Acid ◽  
Interstitial Cells ◽  
Equilibrium Potential ◽  
Inward Currents ◽  
Rabbit Urethra ◽  
Patch Technique ◽  
Dependent Mechanism ◽  
Stimulation Of

Freshly dispersed interstitial cells from the rabbit urethra were studied by using the perforated-patch technique. When cells were voltage clamped at −60 mV and exposed to 10 μM norepinephrine (NE) at 80-s intervals, either large single inward currents or a series of oscillatory inward currents of diminishing amplitude were evoked. These currents were blocked by either phentolamine (1 μM) or prazosin (1 μM), suggesting that the effects of NE were mediated via α1-adrenoceptors. NE-evoked currents were depressed by the blockers of Ca2+-activated Cl− currents, niflumic acid (10 μM), and 9-anthracenecarboxylic acid (9-AC, 1 mM). The reversal potential of the above currents changed in a predictable manner when the Cl− equilibrium potential was altered, again suggesting that they were due to activation of a Cl−conductance. NE-evoked currents were decreased by 10 μM cyclopiazonic acid, suggesting that they were dependent on store-released Ca2+. Inhibition of NE-evoked currents by the phospholipase C inhibitor 2-nitro-4-carboxyphenyl- N, N-diphenylcarbamate (100 μM) suggested that NE releases Ca2+ via an inositol 1,4,5-trisphosphate (IP3)-dependent mechanism. These results support the idea that stimulation of α1-adrenoceptors releases Ca2+ from an IP3-sensitive store, which in turn activates Ca2+-activated Cl− current in freshly dispersed interstitial cells of the rabbit urethra. This elevates slow wave frequency in these cells and may underlie the mechanism responsible for increased urethral tone during nerve stimulation.

scholarly journals Role of IP3 in modulation of spontaneous activity in pacemaker cells of rabbit urethra

2001 ◽  
Vol 280 (5) ◽  
pp. C1349-C1356 ◽  
Author(s):  
G. P. Sergeant ◽  
M. A. Hollywood ◽  
K. D. McCloskey ◽  
N. G. McHale ◽  
K. D. Thornbury
Keyword(s):  
Cyclopiazonic Acid ◽  
Pacemaker Activity ◽  
Pacemaker Cells ◽  
Outward Currents ◽  
Inositol 1,4,5 Trisphosphate ◽  
Inward Currents ◽  
Rabbit Urethra ◽  
Patch Technique ◽  
Spontaneous Transient Outward Currents

Isolated interstitial (“pacemaker”) cells from rabbit urethra were examined using the perforated-patch technique. Under voltage clamp at −60 mV, these cells fired large spontaneous transient inward currents (STICs), averaging −860 pA and >1 s in duration, which could account for urethral pacemaker activity. Spontaneous transient outward currents (STOCs) were also observed and fell into two categories, “fast” (<100 ms in duration) and “slow” (>1 s in duration). The latter were coupled to STICs, suggesting that they shared the same mechanism, while the former occurred independently at faster rates. All of these currents were abolished by cyclopiazonic acid, caffeine, or ryanodine, suggesting that they were activated by Ca2+ release. Whend- myo-inositol 1,4,5-trisphosphate (IP3)-sensitive stores were blocked with 2-aminoethoxydiphenyl borate, the STICs and slow STOCs were abolished, but the fast STOCs remained. In contrast, the fast STOCs were more nifedipine sensitive than the STICs or the slow STOCs. These results suggest that while fast STOCs are mediated by a mechanism similar to STOCs in smooth muscle, STICs and slow STOCs are driven by IP3. These results support the hypothesis that pacemaker activity in the urethra is driven by the IP3-sensitive store.

scholarly journals Activation of Ca2+-activated Cl- current by depolarizing steps in rabbit urethral interstitial cells

2003 ◽  
Vol 285 (2) ◽  
pp. C327-C333 ◽  
Author(s):  
M. A. Hollywood ◽  
G. P. Sergeant ◽  
N. G. McHale ◽  
K. D. Thornbury
Keyword(s):  
Amphotericin B ◽  
Cyclopiazonic Acid ◽  
Interstitial Cells ◽  
Bath Solution ◽  
Perforated Patch ◽  
Inositol 1,4,5 Trisphosphate ◽  
Rabbit Urethra ◽  
Patch Technique ◽  
Aminoethoxydiphenyl Borate

Interstitial cells were isolated from strips of rabbit urethra for study using the amphotericin B perforated-patch technique. Depolarizing steps to -30 mV or greater activated a Ca2+ current ( ICa), followed by a Ca2+-activated Cl- current, and, on stepping back to -80 mV, large Cl- tail currents were observed. Both currents were abolished when the cells were superfused with Ca2+-free bath solution, suggesting that Ca2+ influx was necessary for activation of the Cl- current. The Cl- current was also abolished when Ba2+ was substituted for Ca2+ in the bath or the cell was dialyzed with EGTA (2 mM). The Cl- current was also reduced by cyclopiazonic acid, ryanodine, 2-aminoethoxydiphenyl borate (2-APB), and xestospongin C, suggesting that Ca2+-induced Ca2+ release (CICR) involving both ryanodine and inositol 1,4,5-trisphosphate receptors contributes to its activation.

scholarly journals Na+-K+-Cl− cotransporter (NKCC) maintains the chloride gradient to sustain pacemaker activity in interstitial cells of Cajal

2016 ◽  
Vol 311 (6) ◽  
pp. G1037-G1046 ◽  
Author(s):  
Mei Hong Zhu ◽  
Tae Sik Sung ◽  
Masaaki Kurahashi ◽  
Lauren E. O'Kane ◽  
Kate O'Driscoll ◽  
...  
Keyword(s):  
Slow Wave ◽  
Interstitial Cells Of Cajal ◽  
High Current Density ◽  
Reversal Potential ◽  
Interstitial Cells ◽  
Slow Waves ◽  
Hek293 Cells ◽  
Pacemaker Activity ◽  
Inward Currents ◽  
Cells Of Cajal

Interstitial cells of Cajal (ICC) generate electrical slow waves by coordinated openings of ANO1 channels, a Ca2+-activated Cl− (CaCC) conductance. Efflux of Cl− during slow waves must be significant, as there is high current density during slow-wave currents and slow waves are of sufficient magnitude to depolarize the syncytium of smooth muscle cells and PDGFRα+ cells to which they are electrically coupled. We investigated how the driving force for Cl− current is maintained in ICC. We found robust expression of Slc12a2 (which encodes an Na+-K+-Cl− cotransporter, NKCC1) and immunohistochemical confirmation that NKCC1 is expressed in ICC. With the use of the gramicidin permeabilized-patch technique, which is reported to not disturb [Cl−]i, the reversal potential for spontaneous transient inward currents ( ESTICs) was −10.5 mV. This value corresponds to the peak of slow waves when they are recorded directly from ICC in situ. Inhibition of NKCC1 with bumetanide shifted ESTICs to more negative potentials within a few minutes and reduced pacemaker activity. Bumetanide had no direct effects on ANO1 or CaV3.2 channels expressed in HEK293 cells or L-type Ca2+ currents. Reducing extracellular Cl− to 10 mM shifted ESTICs to positive potentials as predicted by the Nernst equation. The relatively rapid shift in ESTICs when NKCC1 was blocked suggests that significant changes in the transmembrane Cl− gradient occur during the slow-wave cycle, possibly within microdomains formed between endoplasmic reticulum and the plasma membrane in ICC. Recovery of Cl− via NKCC1 might have additional consequences on shaping the waveforms of slow waves via Na+ entry into microdomains.

Inhibitory innervation of colonic smooth muscle cells and interstitial cells of Cajal

1990 ◽  
Vol 68 (3) ◽  
pp. 447-454 ◽  
Author(s):  
Jan D. Huizinga ◽  
Irene Berezin ◽  
Edwin E. Daniel ◽  
Edwin Chow
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Action Potentials ◽  
Interstitial Cells Of Cajal ◽  
Reversal Potential ◽  
Circular Muscle ◽  
Muscle Cells ◽  
Interstitial Cells ◽  
Equilibrium Potential ◽  
Cells Of Cajal

The effect of neural inhibition on the electrical activities of circular and longitudinal colonic smooth muscle was investigated. In addition, a comparative study was carried out between circular muscle preparations with and without the "submucosal" and "myenteric plexus" network of interstitial cells of Cajal (ICC) to study innervation of the "submucosal" ICC and to investigate whether or not the ICC network is an essential intermediary system for inhibitory innervation of smooth muscle cells. Electrical stimulation of intrinsic nerves in the presence of atropine caused inhibitory junction potentials (ijps) throughout the circular and longitudinal muscle layers. The ijp amplitude depended on the membrane potential and not on the location of the muscle cells with respect to the ICC network. Neurally mediated inhibition of the colon resulted in a reduction in amplitude and duration of slow wave type action potentials in circular and abolishment of spike-like action potentials in longitudinal smooth muscle, both resulting in a reduction of contractile activity. With respect to mediation by ICC, the study shows (i) "submucosal" ICC receive direct inhibitory innervation and (ii) circular smooth muscle cells can be directly innervated by inhibitory nerves without ICC as necessary intermediaries. The reversal potential of the ijp in colonic smooth muscle was observed to be approximately −76 mV, close to the estimated potassium equilibrium potential, suggesting that the nerve-mediated hyperpolarization is caused by increased potassium conductance.Key words: enteric nerves, potassium conductance, pacemaker activity, VIP, inhibitory junction potential.

Characterization of a transient outward K+ current with inward rectification in canine ventricular myocytes

1998 ◽  
Vol 274 (3) ◽  
pp. C577-C585 ◽  
Author(s):  
Gui-Rong Li ◽  
Haiying Sun ◽  
Stanley Nattel
Keyword(s):  
Action Potential ◽  
Ventricular Myocytes ◽  
Reversal Potential ◽  
Outward Current ◽  
Equilibrium Potential ◽  
Inward Rectification ◽  
Transient Outward Current ◽  
Membrane Excitability ◽  
Voltage Dependent ◽  
Patch Technique

The threshold potential for the classical depolarization-activated transient outward K+ current and Cl− current is positive to −30 mV. With the whole cell patch technique, a transient outward current was elicited in the presence of 5 mM 4-aminopyridine (4-AP) and 5 μM ryanodine at voltages positive to the K+ equilibrium potential in canine ventricular myocytes. The current was abolished by 200 μM Ba2+ or omission of external K+([Formula: see text]) and showed biexponential inactivation. The current-voltage relation for the peak of the transient outward component showed moderate inward rectification. The transient outward current demonstrated voltage-dependent inactivation (half-inactivation voltage: −43.5 ± 3.2 mV) and rapid, monoexponential recovery from inactivation (time constant: 13.2 ± 2.5 ms). The reversal potential responded to the changes in[Formula: see text] concentration. Action potential clamp revealed two phases of Ba2+-sensitive current during the action potential, including a large early transient component after the upstroke and a later outward component during phase 3 repolarization. The present study demonstrates that depolarization may elicit a Ba2+- and[Formula: see text]-sensitive, 4-AP-insensitive, transient outward current with inward rectification in canine ventricular myocytes. The properties of this K+ current suggest that it may carry a significant early outward current upon depolarization that may play a role in determining membrane excitability and action potential morphology.

scholarly journals Pacemaker activity in urethral interstitial cells is not dependent on capacitative calcium entry

2005 ◽  
Vol 289 (3) ◽  
pp. C625-C632 ◽  
Author(s):  
Eamonn Bradley ◽  
Mark A. Hollywood ◽  
Noel G. McHale ◽  
Keith D. Thornbury ◽  
Gerard P. Sergeant
Keyword(s):  
Spontaneous Activity ◽  
Interstitial Cells ◽  
Calcium Entry ◽  
Pacemaker Activity ◽  
Capacitative Calcium Entry ◽  
Intracellular Ca ◽  
Inward Currents ◽  
Rabbit Urethra ◽  
In Cells

The aim of the present study was to investigate the properties and role of capacitative Ca2+ entry (CCE) in interstitial cells (IC) isolated from the rabbit urethra. Ca2+ entry in IC was larger in cells with depleted intracellular Ca2+ stores compared with controls, consistent with influx via a CCE pathway. The nonselective Ca2+ entry blockers Gd3+ (10 μM), La3+ (10 μM), and Ni2+ (100 μM) reduced CCE by 67% ( n = 14), 65% ( n = 11), and 55% ( n = 9), respectively. These agents did not inhibit Ca2+ entry when stores were not depleted. Conversely, CCE in IC was resistant to SKF-96365 (10 μM), wortmannin (10 μM), and nifedipine (1 μM). Spontaneous transient inward currents were recorded from IC voltage-clamped at −60 mV. These events were not significantly affected by Gd3+ (10 μM) or La3+ (10 μM) and were only slightly decreased in amplitude by 100 μM Ni2+. The results from this study demonstrate that freshly dispersed IC from the rabbit urethra possess a CCE pathway. However, influx via this pathway does not appear to contribute to spontaneous activity in these cells.

Membrane currents evoked by histamine in rabbit basilar artery

1997 ◽  
Vol 272 (2) ◽  
pp. H638-H647 ◽  
Author(s):  
M. Kamouchi ◽  
R. Ogata ◽  
M. Fujishima ◽  
Y. Ito ◽  
K. Kitamura
Keyword(s):  
Membrane Potential ◽  
Basilar Artery ◽  
Reversal Potential ◽  
Outward Current ◽  
Resting Membrane Potential ◽  
Physiological Salt Solution ◽  
Inward Current ◽  
Inward Currents ◽  
Patch Technique ◽  
Isolated Smooth Muscle Cells

The membrane current evoked by histamine in isolated smooth muscle cells from rabbit basilar artery was investigated using the perforated-patch technique. When 10 microM histamine was applied in the bath at a holding potential of -60 mV, an inward current (79.2 +/- 55.8 pA) was transiently activated. An outward current was additionally evoked by 10 microM histamine when the membrane was held at -40 mV or less negative potentials. The outward but not the inward current was completely blocked by 100 nM charybdotoxin. A higher concentration of histamine (30 microM) failed to produce the inward current (3.4 +/- 4.8 pA) when Cl- concentration in the pipette was reduced. The apparent reversal potential of the inward current induced by histamine in physiological salt solution, in high-tetraethylammonium (TEA+) solution (bath), or in low-Cl- solution (pipette) was -6.3 +/- 4.4, -7.5 +/- 4.9, or -45.8 +/- 8.5 mV, respectively. Niflumic acid (100 microM) reversibly blocked the inward current, which was also blocked by 10 microM pyrilamine but not by 10 microM cimetidine. When histamine was continuously applied in the bath, spontaneous transient inward currents were generated. Removal of external Ca2+ or addition of 1 microM nicardipine or 2 mM caffeine reduced the amplitude of the histamine-induced inward current. These results suggest that histamine induces an inward current via H1 receptors at the resting membrane potential, possibly due to activation of Cl- currents. The Cl- inward current might be generated by elevation of intracellular Ca2+ via histamine receptors. The inward current may also contribute to control of the Ca2+ influx via a change in the membrane potential.

scholarly journals Computational modeling of anoctamin 1 calcium-activated chloride channels as pacemaker channels in interstitial cells of Cajal

2014 ◽  
Vol 306 (8) ◽  
pp. G711-G727 ◽  
Author(s):  
Rachel Lees-Green ◽  
Simon J. Gibbons ◽  
Gianrico Farrugia ◽  
James Sneyd ◽  
Leo K. Cheng
Keyword(s):  
Slow Wave ◽  
Interstitial Cells Of Cajal ◽  
Reversal Potential ◽  
Interstitial Cells ◽  
Slow Waves ◽  
Equilibrium Potential ◽  
Pacemaker Activity ◽  
Anoctamin 1 ◽  
Pacemaker Channel ◽  
Cells Of Cajal

Interstitial cells of Cajal (ICC) act as pacemaker cells in the gastrointestinal tract by generating electrical slow waves to regulate rhythmic smooth muscle contractions. Intrinsic Ca2+ oscillations in ICC appear to produce the slow waves by activating pacemaker currents, currently thought to be carried by the Ca2+-activated Cl− channel anoctamin 1 (Ano1). In this article we present a novel model of small intestinal ICC pacemaker activity that incorporates store-operated Ca2+ entry and a new model of Ano1 current. A series of simulations were carried out with the ICC model to investigate current controversies about the reversal potential of the Ano1 Cl− current in ICC and to predict the characteristics of the other ion channels that are necessary to generate slow waves. The model results show that Ano1 is a plausible pacemaker channel when coupled to a store-operated Ca2+ channel but suggest that small cyclical depolarizations may still occur in ICC in Ano1 knockout mice. The results predict that voltage-dependent Ca2+ current is likely to be negligible during the slow wave plateau phase. The model shows that the Cl− equilibrium potential is an important modulator of slow wave morphology, highlighting the need for a better understanding of Cl− dynamics in ICC.

Chloride and cation currents activated by bradykinin in coronary venular endothelial cells

1994 ◽  
Vol 267 (6) ◽  
pp. H2508-H2515
Author(s):  
J. Song ◽  
M. J. Davis
Keyword(s):  
Endothelial Cells ◽  
Time Course ◽  
Reversal Potential ◽  
Disulfonic Acid ◽  
Equilibrium Potential ◽  
K Channel ◽  
Inward Current ◽  
Cation Current ◽  
Inward Currents ◽  
K Current

Bradykinin (BK) is known to activate several types of ion channels in endothelial cells, including a K+ channel and a nonselective cation channel. The predominant BK-activated current in most endothelial cells appears to be an outward, Ca(2+)-activated K+ current. We consistently recorded a rapidly activated, spontaneously inactivated inward current stimulated by BK in bovine coronary venular endothelial cells (CVECs). With the use of a whole cell, perforated patch recording mode, the average magnitude of the current was -293 +/- 38 pA. Simultaneous measurements of current and intracellular Ca2+ concentration ([Ca2+]i) showed that the inward current correlated closely with transient increases in [Ca2+]i due to Ca2+ release from intracellular stores. The current could be blocked by 4,4'-diisothiocyanatostilbene-2, 2'-disulfonic acid (DIDS) but not by La3+, and it persisted in Ca(2+)-free/Na(+)-free solution. When intra- and/or extracellular Cl- concentrations were altered, the reversal potential of the current shifted according to the calculated Cl- -equilibrium potential, indicating that the current was carried primarily by Cl-. Another inward current was also activated by BK. This current was slower to activate, could be blocked by La3+, but was not blocked by DIDS. The time course of the slowly activated current correlated with the plateau phase of the BK-stimulated [Ca2+]i increase, which was similar to the behavior of a nonselective cation current reported previously. We propose that these two currents may contribute to the depolarizations and net inward currents induced by BK in this cell line.

Secretin activates Cl- channels in bile duct epithelial cells through a cAMP-dependent mechanism

1994 ◽  
Vol 266 (4) ◽  
pp. G731-G736 ◽  
Author(s):  
J. M. McGill ◽  
S. Basavappa ◽  
T. W. Gettys ◽  
J. G. Fitz
Keyword(s):  
Bile Duct ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Reversal Potential ◽  
Membrane Current ◽  
Partial Substitution ◽  
Equilibrium Potential ◽  
Resting Membrane Potential ◽  
Patch Clamp Recording ◽  
Dependent Mechanism

Using patch-clamp recording techniques, we assessed the effects of secretin on membrane ion channel activity in isolated rat bile duct epithelial cells. In the whole cell configuration, secretin activated an inward membrane current at -40 mV in 6 of 13 cells, and increased current density from 17 +/- 8 to 98 +/- 33 pA/pF. Secretin-stimulated currents reversed near the equilibrium potential for Cl- and exhibited a linear current-voltage relationship. In the cell-attached configuration, secretin activated low-conductance channels in 73% (11 of 15) of patches. Similar channels were activated by forskolin, suggesting that adenosine 3',5'-cyclic monophosphate (cAMP) is involved as a second messenger. At the resting membrane potential, channels carried inward membrane current and had a slope conductance of 10 +/- 1 pS. In excised patches, addition of purified catalytic subunit of cAMP-dependent protein kinase (protein kinase A) to the cytoplasmic surface activated channels in four of six attempts. With equal Cl- concentrations in bath and pipette, channels had a linear slope conductance of 13 +/- 2 pS and currents reversed near 0 mV. Partial substitution of pipette Cl- with gluconate caused a shift in reversal potential in the direction anticipated for a Cl(-)-selective channel (gluconate to Cl- permeability ratio of 0.21 +/- 0.05, n = 4). Thus in bile duct epithelial cells, exposure to secretin activates low-conductance, Cl(-)-selective channels, probably through a cAMP-dependent mechanism. This likely contributes to secretin-dependent choleresis.

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