scholarly journals Incorporation of RyR2 and Other Ion Channels into Nanopore Based Planar Lipid Bilayers for Low Noise Single Channel Recordings

2010 ◽  
Vol 98 (3) ◽  
pp. 539a-540a
Author(s):  
Prithwish Pal ◽  
Geoffrey A. Barrall ◽  
Ariel L. Escobar ◽  
Melissa A. Poquette ◽  
Patricio Velez ◽  
...  
Keyword(s):  
Ion Channels ◽  
Lipid Bilayers ◽  
Single Channel ◽  
Low Noise ◽  
Planar Lipid Bilayers ◽  
Single Channel Recordings

Ryanodine receptor dysfunction in porcine stunned myocardium

1997 ◽  
Vol 273 (2) ◽  
pp. H796-H804 ◽  
Author(s):  
C. Valdivia ◽  
J. O. Hegge ◽  
R. D. Lasley ◽  
H. H. Valdivia ◽  
R. Mentzer
Keyword(s):  
Coronary Artery ◽  
Lipid Bilayers ◽  
Ryanodine Receptor ◽  
Myocardial Stunning ◽  
Single Channel ◽  
Stunned Myocardium ◽  
Wall Thickening ◽  
Planar Lipid Bilayers ◽  
Open Probability ◽  
Single Channel Recordings

We investigated the effects of myocardial stunning on the function of the two main Ca2+ transport proteins of the sarcoplasmic reticulum (SR), the Ca(2+)-adenosinetriphosphatase and the Ca(2+)-release channel or ryanodine receptor. Regional myocardial stunning was induced in open-chest pigs (n = 6) by a 10-min occlusion of the left anterior descending coronary artery (LAD) and 2 h reperfusion. SR vesicles isolated from the LAD-perfused region (stunned) and the normal left circumflex coronary artery (LC)-perfused region were used to assess the oxalate-supported 45Ca2+ uptake, [3H]ryanodine binding, and single-channel recordings of ryanodine-sensitive Ca(2+)-release channels in planar lipid bilayers. Myocardial stunning decreased LAD systolic wall thickening to 20% of preischemic values. The rate of SR 45Ca2+ uptake in the stunned LAD bed was reduced by 37% compared with that of the normal LC bed (P < 0.05). Stunning was also associated with a 38% reduction in the maximal density of high-affinity [3H]ryanodine binding sites (P < 0.05 vs. normal LC) but had no effect on the dissociation constant. The open probability of ryanodine-sensitive Ca(2+)-release channels determined by single channel recordings in planar lipid bilayers was 26 +/- 2% for control SR (n = 33 channels from 3 animals) and 14 +/- 2% for stunned SR (n = 21 channels; P < 0.05). This depressed activity of SR function observed in postischemic myocardium could be one of the mechanisms underlying myocardial stunning.

scholarly journals Microfabricated Teflon Membranes for Low-Noise Recordings of Ion Channels in Planar Lipid Bilayers

2003 ◽  
Vol 85 (4) ◽  
pp. 2684-2695 ◽  
Author(s):  
Michael Mayer ◽  
Jennah K. Kriebel ◽  
Magdalena T. Tosteson ◽  
George M. Whitesides
Keyword(s):  
Ion Channels ◽  
Lipid Bilayers ◽  
Low Noise ◽  
Planar Lipid Bilayers

scholarly journals Single-channel recordings of RyR1 at microsecond resolution in CMOS-suspended membranes

2018 ◽  
Vol 115 (8) ◽  
pp. E1789-E1798 ◽  
Author(s):  
Andreas J. W. Hartel ◽  
Peijie Ong ◽  
Indra Schroeder ◽  
M. Hunter Giese ◽  
Siddharth Shekar ◽  
...  
Keyword(s):  
Integrated Circuits ◽  
Lipid Bilayers ◽  
Temporal Resolution ◽  
Single Channel ◽  
Low Noise ◽  
Oxide Semiconductor ◽  
Distribution Analysis ◽  
Release Channel ◽  
Single Channel Recordings ◽  
High Bandwidth

Single-channel recordings are widely used to explore functional properties of ion channels. Typically, such recordings are performed at bandwidths of less than 10 kHz because of signal-to-noise considerations, limiting the temporal resolution available for studying fast gating dynamics to greater than 100 µs. Here we present experimental methods that directly integrate suspended lipid bilayers with high-bandwidth, low-noise transimpedance amplifiers based on complementary metal-oxide-semiconductor (CMOS) integrated circuits (IC) technology to achieve bandwidths in excess of 500 kHz and microsecond temporal resolution. We use this CMOS-integrated bilayer system to study the type 1 ryanodine receptor (RyR1), a Ca2+-activated intracellular Ca2+-release channel located on the sarcoplasmic reticulum. We are able to distinguish multiple closed states not evident with lower bandwidth recordings, suggesting the presence of an additional Ca2+ binding site, distinct from the site responsible for activation. An extended beta distribution analysis of our high-bandwidth data can be used to infer closed state flicker events as fast as 35 ns. These events are in the range of single-file ion translocations.

Single K+-channel current measurements from brain synaptosomes in lipid bilayers

1983 ◽  
Vol 245 (1) ◽  
pp. C151-C156 ◽  
Author(s):  
M. T. Nelson ◽  
M. Roudna ◽  
E. Bamberg
Keyword(s):  
Ion Channels ◽  
Lipid Bilayers ◽  
Single Channel ◽  
Mammalian Brain ◽  
K Channel ◽  
Kinetic Properties ◽  
Nerve Terminals ◽  
Planar Lipid Bilayers ◽  
Presynaptic Nerve Terminals ◽  
Single Channel Currents

Ion channels from a rat brain preparation enriched in presynaptic nerve terminals (synaptosomes) were incorporated into planar lipid bilayers. Experiments examined macroscopic (channel-ensemble) currents as well as single-channel currents. Four single-channel conductances (ranging from 10 to 40 pS) were usually observed, each with distinct kinetic properties. All the observed channels selected for K+ over Cl-. These K+ channels may contribute to the resting K+ conductance of brain nerve terminals. Furthermore, this report demonstrates that the properties of ion channels from mammalian brain can be studied in planar lipid bilayers and suggests that this system can be readily extended to many additional investigations on the electrical properties of brain membranes.

Mechanically Stable Lipid Bilayers in Teflon-Coated Silicon Chips for Single-Channel Recordings

2012 ◽  
Vol 4 (1) ◽  
pp. 2-7 ◽  
Author(s):  
Azusa Oshima ◽  
Ayumi Hirano-Iwata ◽  
Tomohiro Nasu ◽  
Yasuo Kimura ◽  
Michio Niwano
Keyword(s):  
Lipid Bilayers ◽  
Single Channel ◽  
Single Channel Recordings ◽  
Silicon Chips

A cloned renal epithelial Na+ channel protein displays stretch activation in planar lipid bilayers

1995 ◽  
Vol 268 (6) ◽  
pp. C1450-C1459 ◽  
Author(s):  
M. S. Awayda ◽  
I. I. Ismailov ◽  
B. K. Berdiev ◽  
D. J. Benos
Keyword(s):  
Hydrostatic Pressure ◽  
Pressure Gradient ◽  
Lipid Bilayers ◽  
Single Channel ◽  
Na Channel ◽  
Planar Lipid Bilayers ◽  
Stretch Activation ◽  
Hydrostatic Pressure Gradient ◽  
Epithelial Na Channel

We have previously cloned a bovine renal epithelial channel homologue (alpha-bENaC) belonging to the epithelial Na+ channel (ENaC) family. With the use of a rabbit nuclease-treated in vitro translation system, mRNA coding for alpha-bENaC was translated and the polypeptide products were reconstituted into liposomes. On incorporation into planar lipid bilayers, in vitro-translated alpha-bENaC protein 1) displayed voltage-independent Na+ channel activity with a single-channel conductance of 40 pS, 2) was mechanosensitive in that the single-channel open probability was maximally activated with a hydrostatic pressure gradient of 0.26 mmHg across the bilayer, 3) was blocked by low concentrations of amiloride [apparent inhibitory constant of amiloride (K(i)amil approximately 150 nM], and 4) was cation selective with a Li+:Na+:K+ permselectivity of 2:1:0.14 under nonstretched conditions. These pharmacological and selectivity characteristics were altered to a lower amiloride affinity (K(i)amil > 25 microM) and a lack of monovalent cation selectivity in the presence of a hydrostatic pressure gradient. This observation of stretch activation (SA) of alpha-bENaC was confirmed in dual electrode recordings of heterologously expressed alpha-bENaC whole cell currents in Xenopus oocytes swelled by the injection of 15 nl of a 100 mM KCl solution. We conclude that alpha-bENaC, and by analogy other ENaCs, represent a novel family of cloned SA channels.

Opening of large-conductance calcium-activated potassium channels by the substituted benzimidazolone NS004

1994 ◽  
Vol 71 (5) ◽  
pp. 1873-1882 ◽  
Author(s):  
M. C. McKay ◽  
S. I. Dworetzky ◽  
N. A. Meanwell ◽  
S. P. Olesen ◽  
P. H. Reinhart ◽  
...  
Keyword(s):  
Rat Brain ◽  
Lipid Bilayers ◽  
Single Channel ◽  
Outward Current ◽  
Calcium Sensitivity ◽  
Bk Channels ◽  
Gh3 Cells ◽  
Xenopus Laevis Oocytes ◽  
Planar Lipid Bilayers ◽  
Whole Cell

1. We used electrophysiological techniques to examine the effects of 5-trifluoromethyl-1-(5-chloro-2-hydroxyphenyl)-1,3-dihydro-2H-benzimidaz ole- 2-one (NS004) on large-conductance calcium-activated potassium (BK) channels. 2. We used recordings from excised membrane patches (cell-attached and inside-out single-channel configurations) and whole-cell patch-clamp recordings to examine the effects of NS004 on single BK channels and whole-cell outward currents, respectively, in rat GH3 clonal pituitary tumor cells. We also tested NS004 on voltage-clamped BK channels isolated from rat brain plasma membrane preparations and reconstituted into planar lipid bilayers. Finally, we used two-electrode voltage-clamp techniques to study the effects of NS004 on currents expressed in Xenopus laevis oocytes by the recently described Slo BK clone from Drosophila. 3. In GH3 cells and in Xenopus oocytes expressing the Slo gene product NS004 produced an increase in an iberiotoxin- or tetraethylammonium-sensitive whole-cell outward current, respectively. NS004 produced a significant increase in the activity of single GH3 cell BK channels and rat brain BK channels reconstituted into planar lipid bilayers. In both systems this was characterized by an increase in channel mean open time, a decrease in interburst interval, and an apparent increase in channel voltage/calcium sensitivity. 4. These data indicate that NS004 could be useful for investigating the biophysical and molecular properties of BK channels and for determining the functional consequences of the opening of BK channels.

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