scholarly journals Expression and function of KV2-containing channels in human urinary bladder smooth muscle

2012 ◽  
Vol 302 (11) ◽  
pp. C1599-C1608 ◽  
Author(s):  
Kiril L. Hristov ◽  
Muyan Chen ◽  
Serge A. Y. Afeli ◽  
Qiuping Cheng ◽  
Eric S. Rovner ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Patch Clamp ◽  
Electrical Field Stimulation ◽  
Bladder Smooth Muscle ◽  
Rt Pcr ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp

The functional role of the voltage-gated K+ (KV) channels in human detrusor smooth muscle (DSM) is largely unexplored. Here, we provide molecular, electrophysiological, and functional evidence for the expression of KV2.1, KV2.2, and the electrically silent KV9.3 subunits in human DSM. Stromatoxin-1 (ScTx1), a selective inhibitor of KV2.1, KV2.2, and KV4.2 homotetrameric channels and of KV2.1/9.3 heterotetrameric channels, was used to examine the role of these channels in human DSM function. Human DSM tissues were obtained during open bladder surgeries from patients without a history of overactive bladder. Freshly isolated human DSM cells were studied using RT-PCR, immunocytochemistry, live-cell Ca2+ imaging, and the perforated whole cell patch-clamp technique. Isometric DSM tension recordings of human DSM isolated strips were conducted using tissue baths. RT-PCR experiments showed mRNA expression of KV2.1, KV2.2, and KV9.3 (but not KV4.2) channel subunits in human isolated DSM cells. KV2.1 and KV2.2 protein expression was confirmed by Western blot analysis and immunocytochemistry. Perforated whole cell patch-clamp experiments revealed that ScTx1 (100 nM) inhibited the amplitude of the voltage step-induced KV current in freshly isolated human DSM cells. ScTx1 (100 nM) significantly increased the intracellular Ca2+ level in DSM cells. In human DSM isolated strips, ScTx1 (100 nM) increased the spontaneous phasic contraction amplitude and muscle force, and enhanced the amplitude of the electrical field stimulation-induced contractions within the range of 3.5–30 Hz stimulation frequencies. These findings reveal that ScTx1-sensitive KV2-containing channels are key regulators of human DSM excitability and contractility and may represent new targets for pharmacological or genetic intervention for bladder dysfunction.

scholarly journals Baro-excited neurons in the caudal ventrolateral medulla (CVLM) recorded using the whole-cell patch-clamp technique

2011 ◽  
Vol 35 (5) ◽  
pp. 500-506 ◽  
Author(s):  
Naoki Oshima ◽  
Hiroo Kumagai ◽  
Kamon Iigaya ◽  
Hiroshi Onimaru ◽  
Akira Kawai ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Ventrolateral Medulla ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Caudal Ventrolateral Medulla ◽  
Clamp Technique ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp

scholarly journals Neutrophil Secretion Induced by an Intracellular Ca2+Rise and Followed by Whole-Cell Patch-Clamp Recordings Occurs Without any Selective Mobilization of Different Granule Populations

2006 ◽  
Vol 2006 ◽  
pp. 1-7 ◽  
Author(s):  
Daniel Granfeldt ◽  
Olle Harbecke ◽  
Åse Björstad ◽  
Anna Karlsson ◽  
Claes Dahlgren
Keyword(s):  
Patch Clamp ◽  
Human Neutrophils ◽  
Lag Phase ◽  
Measuring Time ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
High Concentrations ◽  
Kinetics Of

We have investigated calcium-induced secretion in human neutrophils, using a whole-cell patch-clamp technique. Mobilization of subcellular granules to the cell membrane was followed as the change in membrane capacitance (△Cm). Both the magnitude and the kinetics of the response differed between low and high concentrations of Ca2+. A sustained secretion following a short lag phase was induced by high concentrations of Ca2+(100μM and higher). A stable plateau was reached after 5–7 minutes at△Cmvalues corresponding to values expected after all specific as well as azurophil granules have been mobilized. Capacitance values of the same magnitude could be obtained also at lower Ca2+concentrations, but typically no stable plateau was reached within the measuring time. In contrast to previous studies, we were unable to detect any pattern of secretion corresponding to a distinct submaximal response or selective mobilization of granule subsets specified by their Ca2+-sensitivity.

GABA-Mimetic Actions of Withania somnifera on Substantia Gelatinosa Neurons of the Trigeminal Subnucleus Caudalis in Mice

2013 ◽  
Vol 41 (05) ◽  
pp. 1043-1051 ◽  
Author(s):  
Hua Yin ◽  
Dong Hyu Cho ◽  
Soo Joung Park ◽  
Seong Kyu Han
Keyword(s):  
Patch Clamp ◽  
Receptor Antagonist ◽  
Withania Somnifera ◽  
Substantia Gelatinosa ◽  
Patch Clamp Technique ◽  
Pipette Solution ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
Inward Currents

The plant Withania somnifera (WS), also known as Ashwagandha, has been used widely in traditional medicine systems in India and Nepal (Ayurveda), and has been accepted to cure various ailments. In this study, the whole-cell patch clamp technique was performed to examine the mechanism of action of WS on the SG neurons of the Vc from mouse brainstem slices. In whole-cell patch clamp mode, methanol extract of Withania somnifera (mWS) induced short-lived and repeatable inward currents in all SG neurons tested (31.3±8.51 pA, n = 7) using a high chloride pipette solution. The mWS-induced inward currents were concentration dependent and maintained in the presence of tetrodotoxin (TTX), a voltage gated Na + channel blocker, CNQX, a non-NMDA glutamate receptor antagonist, AP5, an NMDA receptor antagonist and strychnine, a glycine receptor antagonist. The mWS induced currents were blocked by picrotoxin, a GABAA receptor antagonist. These results show that mWS has an inhibitory effects on SG neurons of the Vc through GABAA receptor-mediated activation of chloride ion channels, indicating that mWS contains compounds with sedative effects on the central nervous system. These results also suggest that mWS may be a potential target for modulating orofacial pain processing.

Properties of Ca2+-mediated inactivation of L-type Ca channel in smooth muscle cells of the guinea-pig urinary bladder

1995 ◽  
Vol 73 (1) ◽  
pp. 27-35 ◽  
Author(s):  
M. Yoshino ◽  
Y. Matsufuji ◽  
H. Yabu
Keyword(s):  
Smooth Muscle ◽  
Urinary Bladder ◽  
Patch Clamp ◽  
Double Pulse ◽  
Ca Channel ◽  
Time Constants ◽  
Whole Cell ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp ◽  
Pulse Experiment

The properties of Ca2+-mediated inactivation as revealed by a conventional double-pulse protocol were examined by using the whole-cell patch clamp technique. A U-shaped relationship between the conditioning potential and the Ca2+ current (ICa) inactivation was observed, with a maximum inactivation of 52 ± 4% (n = 5) at 10 mV with 0.5 mM EGTA in the patch pipettes. The maximum inactivation was reduced significantly, to 31 ± 5.7% (n = 12) and 32 ± 7.0% (n = 5), when a high concentration of EGTA (20 mM) or a more efficient Ca2+ chelator, BAPTA, was included in the patch pipettes, respectively. The same double-pulse protocol was applied under conditions where the stored Ca2+ was depleted by using caffeine or the stored Ca2+ release function was blocked by using ryanodine or procaine and heparin. No significant difference in the maximum ICa inactivation before (45%) and after (50%) application of 10 mM caffeine was observed. The maximum ICa inactivations of 48 ± 3.2% (n = 4) and 52 ± 8.4% (n = 6) were still observed after treatment of the cell with ryanodine (20 μM) or loading 10 mM procaine and 1 mg/mL heparin in the patch pipettes, respectively. These results suggest that Ca2+ mobilization from an internal Ca2+ store is not essential for the Ca2+-mediated inactivation observed in the double-pulse experiment, rather influx of Ca2+ through a voltage-dependent Ca channel seems to be important for ICa inactivation. Recovery from Ca2+-mediated inactivation could be fitted by the sum of two exponentials; the time constants of the fast and slow components were 210 and 810 ms, respectively, at a holding potential of −80 mV. By lowering the holding potential to −30 from −80 mV, the time constants of both fast and slow components were increased to 740 and 4820 ms, respectively, suggesting that the recovery of Ca2+ channels from Ca2+-mediated inactivation is voltage dependent.Key words: urinary bladder, smooth muscle, whole-cell patch clamp, Ca2+ current, double-pulse protocol, Ca2+-mediated inactivation.

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